Compositions for prevention of adhesions and other barrier applications

ABSTRACT

A method has been developed of preventing or limiting formation of adhesions by administering to a site in need thereof, in the absence of or after bleeding or leakage of fluid has been substantially stopped, a self-assembling material which forms a barrier to formation of adhesions. In certain embodiments, the self assembling materials are peptidomimetics, nucleotidomimetics, di- and triblock copolymers, N-alkylacrylamides, or dendrimers. These materials are also useful in a method for regeneration or repair of tissue or cells forming tissue.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional application of U.S. Ser. No.14/885,673, filed on Oct. 16, 2015, which is a divisional application ofU.S. Ser. No. 11/740,284, filed on Apr. 25, 2007, now U.S. Pat. No.9,612,005, issued Oct. 20, 2015, which claims the benefit of andpriority to U.S. Ser. No. 60/745,601, filed on Apr. 25, 2006.

REFERENCE TO SEQUENCE LISTING

The sequence listing submitted Sep. 7, 2007 as a text file named“CNS_102_ST25.txt,” created on Jul. 6, 2007, and having a size of 91,000bytes is hereby incorporated by reference pursuant to 37 C.F.R. §1.52(e)(5).

FIELD OF THE INVENTION

The present invention is generally in the field of formulations forapplication to tissues for prevention of adhesions and other barrierapplications.

BACKGROUND OF THE INVENTION

Adhesions may be present at birth (congenital) or may form afterabdominal surgery or inflammation. Most form after surgery. Adhesionsare more common after procedures on the colon, appendix, or uterus thanafter surgery on the stomach, gall bladder, or pancreas. The risk ofdeveloping adhesions increases with the passage of time after thesurgery.

Abdominal adhesions are bands of fibrous scar tissue that form on organsin the abdomen, causing the organs to stick to one another or to thewall of the abdomen. Intestinal adhesions are bands of fibrous tissuethat connect the loops of the intestines to each other, or theintestines to other abdominal organs, or the intestines to the abdominalwall. These bands can pull sections of the intestines out of place andmay block passage of food. In people living in developed countries, thisscar tissue most commonly develops after abdominal surgery, in whichorgans are handled by the surgical team and are shifted temporarily fromtheir normal positions. It can also form in people who developperitonitis, an infection that has spread to the membrane that coversthe abdominal organs. Peritonitis commonly occurs after appendicitis orother abdominal infections. Another cause of adhesions is endometriosis,an inflammatory condition that affects some women and may involve theabdomen and serious abdominal trauma, including cesarean sections.

Adhesions are a major cause of intestinal obstruction. If the adhesionscause partial or complete obstruction of the intestines, the symptomsexhibited depend on the degree and the location of the obstruction. Theyinclude cramps, abdominal pain, vomiting, bloating, an inability to passgas, and constipation. In a small number of people who have adhesions,however, the fibrous bands of scar tissue block the intestines eithercompletely or partially. This blockage is called a bowel obstruction,and leads to death in about 5% of cases. Sometimes, an area of intestinethat is affected by adhesions can become blocked then unblocked, causingsymptoms to come and go. In about 10% of small-bowel obstructions, aportion of the bowel twists tightly around a band of adhesions. Thiscuts off the normal blood supply to the twisted bowel, a disorder knownas strangulation, causing that section of bowel to die. When thisemergency happens, the person must undergo surgery immediately. Thedeath rate is as high as 37% in people who develop strangulation.

Percutaneous epidural adhesiolysis and spinal endoscopic adhesiolysisare interventional pain management techniques used to treat patientswith refractory low back pain due to epidural scarring. Standardepidural steroid injections are often ineffective, especially inpatients with prior back surgery. Adhesions in the epidural space canprevent the flow of medicine to the target area; lysis of theseadhesions can improve the delivery of medication to the affected areas,potentially improving the therapeutic efficacy of the injectedmedications. Prevention of such adhesions would be more preferable.

Many different materials have been tried as a means of preventingadhesions. Most of these are hydrogels that are applied as solutions atthe time of surgery. Efficacy of these materials has varied due to rapiddegradation and/or failure to form a sufficiently thick barrier. Othersmaterials work only in combination with anti-proliferative drugs. Noneof these materials has been shown to be effective work in a highly fluidenvironment, which usually is present during surgery due to bleeding andleakage of other bodily fluids.

It is therefore an object of the present invention to provide methodsand compositions for preventing or minimizing adhesions and for otherbarrier applications which can be applied to tissues or cells which arebleeding or in the presence of fluids.

It is another object of the present invention to provide such acomposition that can be formulated as a bandage, spray, coating, orpowder.

It is a still further object of the present invention to provide acomposition that can be used to retain fluids or cells but issufficiently clear to allow a physician to see and work through thematerial.

BRIEF SUMMARY OF THE INVENTION

Compositions including materials which self-assemble under physiologicalconditions are formulated for application to tissues for prevention ofadhesions or other barrier applications, such as minimizingcontamination or infection, limiting spread of metastasis followingcancer surgery, or for delivery of a therapeutic, diagnostic orprophylactic agent in a confined area, after bleeding or fluid leakagehas been substantially stopped. In one embodiment, the self-assemblingmaterial comprises peptides having a sequence of amino acid residuesconforming to one or more of Formulas I-IV:((Xaa^(o)′−Xaa+)x(Xaane1_Xaa−)y)n (I);((Xaa′_Xaa)x(Xaa^(o)e^(o)−Xaa+)y)n (II); ((Xaa+−Xaa″e1)x(Xaa−Xaa″)y)n(III); and ((Xaa−XaaQe7)x(Xaa+Xaa1e″)y)n (IV), where Xaan′ represents anamino acid residue having a neutral charge; Xaa+ represents an aminoacid residue having a positive charge; Xaa represents an amino acidresidue having a negative charge; x and y are integers having a value of1, 2 or 4, independently; and n is an integer having a value of 1-5. Inanother embodiment, the self assembling materials are peptidomimetics,nucleotidomimetics, di- and triblock copolymers, N-alkylacrylamides, ordendrimers. These materials are also useful in a method for regenerationor repair of tissue or cells forming tissue.

The concentration of the self-assembling materials in any givenformulation can vary and can be between approximately 0.1% and 99%,inclusive, preferably between 0.1% and 10%. In one embodiment, theconcentration of the self-assembling materials (e.g., in a liquidformulation) can be approximately 0.1-3.0% (1-30 mg/ml) (e.g., 0.1-1.0%;1.0-2.0%; 2.0-3.0% or 1.0-3.0%). The concentration of self-assemblingmaterials can be higher in stock solutions and in solid (e.g., powdered)formulations. Solid preparations may have a concentration ofself-assembling materials approaching 100% (e.g., the concentration ofself-assembling materials can be 95, 96, 97, 98, 99% or more (e.g.,99.99%) of the composition). Whether in liquid or solid form, thematerials can be brought to the desired concentration prior to use byaddition of a pharmaceutically acceptable diluent (e.g. deionizedwater), fillers, or oil. The formulations may include a pharmaceuticallyacceptable carrier or therapeutic, prophylactic or diagnostic agents.These include, but are not limited to, anti-inflammatories, vasoactiveagents, anti-infectives, anesthetics, growth factors, and/or cells.Metals may be added as chelators or to further decrease adhesion.

The formulation can be administered as appropriate for treatment of oneor more disorders or conditions, such as those noted above. For example,the formulation may be applied after repair of an injury or duringsurgery of the lung, eye or dura, or following an epidural or spinaltap, to prevent or minimize formation of adhesions. The formulation maybe administered to a burn or ulcer, after bleeding or fluid loss isstopped.

The formulation may be applied as a hydrogel, laminate including oil, orspray. In one embodiment, the formulation is provided as a dry orlyophilized powder which can be administered directly as a powder or atablet, disc, or wafer which hydrates at the site of application, orsuspended or dissolved in a liquid, most preferably aqueous, and appliedas a spray, paint, injection or a hydrogel including a material such aschitin, collagen, alginate, or synthetic polymer. In the preferredembodiment, the material is provided in combination with an oil, andforms a laminate. In yet another embodiment, the formulation is providedin a bandage, foam or matrix, in which the materials may be dispersed orabsorbed. The formulation could also be in the form of sutures, tape, oradhesive. The liquid formulations may be provided in a syringe orpipette having a barrel containing a composition includingself-assembling materials and a means for expelling the composition froman open tip of the syringe or pipette (e.g., a plunger or bulb). Thesyringe may consist of one or more compartments, so that mixing of theself-assembling materials with one or more other agents occurs at thetime of application. The compartments may also contain excipients suchas a material forming a hydrogel or adhesive in one compartment and theself-assembling materials in the other compartment. In anotherembodiment, one compartment may contain lyophilized or particles ofself-assembling materials, and another compartment may contain solutionto dissolve or hydrate the materials, or mixed with other powders fordry application. The liquid and powder compositions are stable,preferably for a period greater than one year, more preferably greaterthan two years and most preferably greater than three years.

One or more of the compositions described herein can be assembled inkits, together with instructions for use. For example, the kits caninclude a biocompatible composition including self-assembling materials(or a concentrated solution or powdered formulation thereof, togetherwith a diluent) and a vasoconstrictor, a coloring agent, or an analgesicor anesthetic agent and instructions for their combination (if notalready combined) and use (e.g., dilution and administration). The kitscan further include one or more of the additional agents describedherein. These agents can be present within the self assemblingcomposition or packaged separately, and they can include one or moretypes of biological cells, an antibiotic or other therapeutic, collagen,an anti-inflammatory agent, a growth factor, or a nutrient. The kit mayalso include one or more of a syringe (e.g., a barrel syringe or a bulbsyringe), a needle, a pipette, gauze, sponges, cotton, swabs, a bandage,a disinfectant, surgical thread, scissors, a scalpel, a sterile fluid, aspray canister, including those in which a liquid solution is sprayedthrough a simple hand pump, a sterile container, or disposable gloves.

DETAILED DESCRIPTION OF THE INVENTION

I. Formulations

“Biocompatible”, as used herein, refers to compatibility with livingtissue or a living system by not being toxic, injurious, orphysiologically reactive and not causing immunological rejection.

“Complementary” means having the capability of forming ionic or hydrogenbonding interactions between hydrophilic residues from adjacent peptidesin a structure. Each hydrophilic reside in a peptide either hydrogenbonds or ionically pairs with a hydrophilic residue on an adjacentpeptide, or is exposed to solvent. Pairing may also involve van derWaals forces.

“Effective amount”, in reference to an active agent such as aself-assembling peptide or biomolecule, pharmaceutical agent, etc.refers to the amount necessary to elicit a desired biological response.As will be appreciated by those of ordinary skill in this art, theeffective amount of an agent may vary depending on such factors as thedesired biological endpoint, the agent to be delivered, the nature ofthe site to which the agent is delivered, the nature of the conditionsfor which the agent is administered, etc. For example, the effectiveamount of a composition for treatment of diabetic retinopathy may be anamount sufficient to promote recovery to a greater extent than wouldoccur in the absence of the composition.

“Hemostasis” refers to the cessation of bleeding.

“Preventing” refers to causing a condition, state, or disease, orsymptom or manifestation of such, or worsening of the severity of such,not to occur. Preventing includes reducing the risk that a condition,state, or disease, or symptom or manifestation of such, or worsening ofthe severity of such, will occur.

“Repair”, as sued in reference to the repair of tissue in variousembodiments of the invention, may include any aspect of anatomical orfunctional restoration of the condition of the tissue prior to aninjury, deterioration, or other damage. For example, it may includerestoration of physical continuity between portions of tissue that wereseparated by injury, deterioration, or other damage. Preferably suchrestoration of physical continuity includes reposition or reconnectionof the portions of tissue without appreciable separation by tissue of atype that was not present prior to the injury, such as scar tissue.Repair may, but need not, include growth or development of new tissue.“Repair” and “Healing” are used interchangeably herein.

II. Self-Assembling Materials

A. Self-Assembling Peptides

In one embodiment, the self-assembling material is a self-assemblingpeptide. The term “peptide,” as used herein includes “polypeptide,”“oligopeptide,” and “protein,” and refers to a chain of at least twoα-amino acid residues linked together by covalent bonds (e.g., peptidebonds). Useful peptides can vary in length so long as they retain theability to self-assemble to an extent useful for one or more of thepurposes described herein. The number of amino acid residues in thepeptide may range from as few as two α-amino acid residues to about 200residues. Typically, peptides which self-assemble have from about 6 toabout 200 residues, preferably from about 6 to about 64 residues, morepreferably from about 8 to about 36 residues, most preferably from about8 to about 24 residues. The peptides can be at least eight amino acidsin length (e.g., eight or 10 amino acids), at least 12 amino acids inlength (e.g., 12 or 14 amino acids), or at least 16 amino acids inlength (e.g., 16, 18, 20, 22, or 24 amino acids). Peptides that are lessthan 100 amino acid residues long, more preferably less thanapproximately 50 amino acids in length, may assemble more readily. Inone embodiment, the peptide has from about 8 to about 16 residues. Inanother embodiment, the peptide has from about 12 to about 20 residues.In yet another embodiment, the peptide has from about 16 to about 20residues. “Peptide” may refer to an individual peptide or to acollection of peptides having the same or different sequences, any ofwhich may contain naturally occurring α-amino acid residues,non-naturally occurring α-amino acid residues, and combinations thereof.α-Amino acid analogs are also known in the art and may alternatively beemployed. In particular, D-α-amino acid residues may be used.

In addition, one or more of the amino acid residues in a self-assemblingpeptide can be altered or derivatized by the addition of one or morechemical entities including, but not limited to, acyl groups,carbohydrate groups, carbohydrate chains, phosphate groups, farnesylgroups, isofarnesyl groups, fatty acid groups, or a linker which allowsfor conjugation or functionalization of the peptide.

While the sequences of the peptides can vary, useful sequences includethose that convey an amphiphilic nature to the peptides (e.g., thepeptides can contain approximately equal numbers of hydrophobic andhydrophilic amino acid residues), and the peptides can be complementaryand structurally compatible. Complementary peptides have the ability toform ionic or hydrogen bonds between residues (e.g., hydrophilicresidues) on adjacent peptides in a structure. For example, one or morehydrophilic residues in a peptide can either hydrogen bond or ionicallypair with one or more hydrophilic residues on an adjacent peptide.Unpaired residues can interact (e.g. form hydrogen bonds, etc.,) withthe solvent. Peptide-peptide interactions may also involve van der Waalsforces and/or forces that do not constitute covalent bonds. The peptidesare structurally compatible when they are capable of maintaining asufficiently constant intrapeptide distance to allow self-assembly andstructure formation. The intrapeptide distance can vary. “Intrapeptidedistance”, as used herein, refers to the average of a representativenumber of distances between adjacent amino acid residues. In oneembodiment, the intrapeptide distance is less than about 4 angstroms,preferably less than about 3, more preferably less than about 2angstroms, and most preferably less than about 1 angstrom. Theintrapeptide distance may be larger than this, however. These distancescan be calculated based on molecular modeling or based on a simplifiedprocedure described in U.S. Pat. No. 5,670,483 to Zhang et al.

Where self-assembling peptides are used, it is thought that their sidechains (or R groups) partition into two faces, a polar face withpositively and/or negatively charged ionic side chains, and a nonpolarface with side chains that are considered neutral or uncharged atphysiological pH (e.g., the side chain of an alanine residue or residueshaving other hydrophobic groups). The positively charged and negativelycharged amino acid residues on the polar face of one peptide can formcomplementary ionic pairs with oppositely charged residues of anotherpeptide. These peptides may therefore be called ionic,self-complementary peptides. If the ionic residues alternate with onepositively and one negatively charged residue on the polar face (− + − +−+ − +), the peptides may be described as “modulus I;” if the ionicresidues alternate with two positively and two negatively chargedresidues (− − + + − − ++) on the polar face, the peptides are describedas “modulus II;” if the ionic residues alternate with three positivelyand three negatively charged residues (+++−−−+++−−−) on the polar face,the peptides are describe as “modulus III;” if the ionic residuesalternate with four positively and four negatively charged residues(++++−−−++++−−−−) on the polar face, they are described as “modulus IV.”A peptide having four repeating units of the sequence EAKA (SEQ ID NO:2) may be designated EAKA16-I, and peptides having other sequences maybe described by the same convention.

Peptide-based structures can be formed of heterogeneous mixtures ofpeptides (i.e., mixtures containing more than one type of peptideconforming to a given formula or to two or more of the formulas). Insome embodiments, each of the types of peptides in the mixture are ableto self-assemble alone. In other embodiments, one or more of each typeof peptide would not, alone, self-assemble but the combination ofheterogeneous peptides may self-assemble (i.e., peptides in the mixtureare complementary and structurally compatible with each other). Thus,either a homogeneous mixture of self-complementary and self-compatiblepeptides of the same sequence or containing the same repeating subunit,or a heterogeneous mixture of different peptides which are complementaryand structurally compatible to each other, can be used. The compositionsdescribed herein regardless of the precise form (e.g., whether in aliquid form or molded) and regardless of the overall compositions (e.g.,whether combined with another agent, contained within a device, orpackaged in a kit) can include a mixture of one or more peptide chains.

Either or both ends of a given peptide can be modified. For example, thecarboxyl and/or amino groups of the carboxyl- and amino-terminalresidues, respectively can be protected or not protected. The charge ata terminus can also be modified. For example, a group or radical such asan acyl group (RCO—, where R is an organic group (e.g., an acetyl group(CH₃CO—)) can be present at the N-terminus of a peptide to neutralize an“extra” positive charge that may otherwise be present (e.g., a chargenot resulting from the side chain of the N-terminal amino acid).Similarly, a group such as an amine group (NH₂) can be used toneutralize an “extra” negative charge that may otherwise be present atthe C-terminus (e.g., a charge not resulting from the side chain of theC-terminal amino acid residue). Where an amine is used, the C-terminuswould bear an amide (—CONH₂). The neutralization of charges on aterminus may facilitate self-assembly. One of ordinary skill in the artwill be able to select other suitable groups.

Useful peptides can also be branched, in which case they will contain atleast two amino acid polymers, each of which consists of at least threeamino acid residues joined by peptide bonds. The two amino acid polymersmay be linked by a bond other than a peptide bond.

The structures described herein can be formed through self-assembly ofthe peptides described in U.S. Pat. Nos. 5,670,483; 5,955,343;6,548,630; and 6,800,481 to Zhang et al.; Holmes et al., Proc. Natl.Acad. Sci. USA, 97:6728-6733 (2000); Zhang et al., Proc. Natl. Acad.Sci. USA, 90:3334-3338 (1993); Zhang et al., Biomaterials, 16:1385-1393(1995); Caplan et al., Biomaterials, 23:219-227 (2002); Leon et al., J.Biomater. Sci. Polym. Ed., 9:297-312 (1998); and Caplan et al.,Biomacromolecules, 1:627-631 (2000).

Self-assembling peptides containing alternating hydrophobic andhydrophilic amino residues can be used. Examples of representativehydrophobic and hydrophilic peptides are listed in Table 1.

TABLE 1 Representative Self-Assembling Peptides No. Sequence (N → C) 1.n-SGSGSGSGSGSGSGSG-c (SEQ ID NO: 3) 2. n-SASASASASASASASA-c(SEQ ID NO: 4) 3. n-SVSVSVSVSVSVSVSV-c (SEQ ID NO: 5) 4.n-SLSLSLSLSLSLSLSL-c (SEQ ID NO: 6 5. n-SISISISISISISISI-c(SEQ ID NO: 7) 6. n-SMSMSMSMSMSMSMSM-c (SEQ ID NO: 8) 7.n-SFSFSFSFSFSFSFSF-c (SEQ ID NO: 9) 8. n-SWSWSWSWSWSWSWSW-c(SEQ ID NO: 10) 9. n-SPSPSPSPSPSPSPSP-c (SEQ ID NO: 11) 10.n-TGTGTGTGTGTGTGTG-c (SEQ ID NO: 12) 11. n-TATATATATATATATA-c(SEQ ID NO: 13) 12. n-TVTVTVTVTVTVTVTV-c (SEQ ID NO: 14) 13.n-TLTLTLTLTLTLTLTL-c (SEQ ID NO: 15) 14. n-TITITITITITITITI-c(SEQ ID NO: 16) 15. n-TMTMTMTMTMTMTMTM-c (SEQ ID NO: 17) 16.n-TFTFTFTFTFTFTFTF-c (SEQ ID NO: 18) 17. n-TWTWTWTWTWTWTWTW-c(SEQ ID NO: 19) 18. n-TPTPTPTPTPTPTPTP-c (SEQ ID NO: 20) 19.n-CGCGCGCGCGCGCGCG-c (SEQ ID NO: 21) 20. n-CACACACACACACACA-c(SEQ ID NO: 22) 21. n-CVCVCVCVCVCVCVCV-c (SEQ ID NO: 23) 22.n-CLCLCLCLCLCLCLCL-c (SEQ ID NO: 24) 23. n-CICICICICICICICI-c(SEQ ID NO: 25) 24. n-CMCMCMCMCMCMCMCM-c (SEQ ID NO: 26) 25.n-CFCFCFCFCFCFCFCF-c (SEQ ID NO: 27) 26. n-CWCWCWCWCWCWCWC-c(SEQ ID NO: 28) 27. n-CPCPCPCPCPCPCPCP-c (SEQ ID NO: 29) 28.n-YGYGYGYGYGYGYGYG-c (SEQ ID NO: 30) 29. n-YAYAYAYAYAYAYAYA-c(SEQ ID NO: 31) 30. n-YVYVYVYVYVYVYVYV-c (SEQ ID NO: 32) 31.n-YLYLYLYLYLYLYLYL-c (SEQ ID NO: 33) 32. n-YIYIYIYIYIYIYIYI-c(SEQ ID NO: 34) 33. n-YMYMYMYMYMYMYMYM-c (SEQ ID NO: 35) 34.n-YFYFYFYFYFYFYFYF-c (SEQ ID NO: 36) 35. n-YWYWYWYWYWYWYWYW-c(SEQ ID NO: 37) 36. n-YPYPYPYPYPYPYPYP-c (SEQ ID NO: 38) 37.n-NGNGNGNGNGNGNGNG-c (SEQ ID NO: 39) 38. n-NANANANANANANANA-c(SEQ ID NO: 40) 39. n-NVNVNVNVNVNVNVNV-c (SEQ ID NO: 41) 40.n-NLNLNLNLNLNLNLNL-c (SEQ ID NO: 42) 41. n-NINININININININI-c(SEQ ID NO: 43) 42. n-NMNMNMNMNMNMNMNM-c (SEQ ID NO: 44) 43.n-NFNFNFNFNFNFNFNF-c (SEQ ID NO: 45) 44. n-NWNWNWNWNWNWNWNW-c(SEQ ID NO: 46) 45. n-NPNPNPNPNPNPNPNP-c (SEQ ID NO: 47) 46.n-QGQGQGQGQGQGQGQG-c (SEQ ID NO: 48) 47. n-QAQAQAQAQAQAQAQA-c(SEQ ID NO: 49) 48. n-QVQVQVQVQVQVQVQV-c (SEQ ID NO: 50) 49.n-QLQLQLQLQLQLQLQL-c (SEQ ID NO: 51) 50. n-QIQIQIQIQIQIQIQI-c(SEQ ID NO: 52) 51. n-QMQMQMQMQMQMQMQM-c (SEQ ID NO: 53) 52.n-QFQFQFQFQFQFQFQF-c (SEQ ID NO: 54) 53. n-QWQWQWQWQWQWQWQW-c(SEQ ID NO: 55) 54. n-QPQPQPQPQPQPQPQP-c (SEQ ID NO: 56)

Other useful self-assembling peptides can be generated, for example,which differ from those exemplified by a single amino acid residue or bymultiple amino acid residues (e.g., by inclusion or exclusion of arepeating quartet). For example, one or more cysteine residues may beincorporated into the peptides, and these residues may bond with oneanother through the formation of disulfide bonds. Structures bonded inthis manner may have increased mechanical strength relative tostructures made with comparable peptides that do not include cysteineresidues and thus are unable to form disulfide bonds.

The amino acid residues in the self-assembling peptides can be naturallyoccurring or non-naturally occurring amino acid residues. Naturallyoccurring amino acids can include amino acid residues encoded by thestandard genetic code as well as non-standard amino acids (e.g., aminoacids having the D-configuration instead of the L-configuration), aswell as those amino acids that can be formed by modifications ofstandard amino acids (e.g. pyrolysine or selenocysteine). Non-naturallyoccurring amino acids are not been found in nature, but can beincorporated into a peptide chain. Suitable non-naturally occurringamino acids include, but are not limited to,D-alloisoleucine(2R,3S)-2-amino-3-methylpentanoic acid, L-cyclopentylglycine (S)-2-amino-2-cyclopentyl acetic acid. Other examples ofnon-naturally occurring amino acids can be found in textbooks or on theworldwide web (e.g., a site is maintained by the California Institute ofTechnology which displays structures of non-natural amino acids thathave been successfully incorporated into functional proteins).Non-natural amino acid residues and amino acid derivatives described inU.S. Patent Application Publication No. 2004/0204561 to Ellison.

Self-assembling peptides can be chemically synthesized or purified fromnatural or recombinantly-produced sources by methods well known in theart. For example, peptides can be synthesized using standard f-mocchemistry and purified using high pressure liquid chromatography (HPLC).

Self-complementary peptides such as EAKA16-I, RADA16-I (SEQ ID NO: 1),RAEA16-I, and KADA16-I are described in Zhang, S., et al. ((1999)Peptide self-assembly in functional polymer science and engineering.Reactive & Functional Polymers, 41, 91-102). The self-assemblingpeptides comprise a sequence of amino acid residues conforming to one ormore of Formulas I-IV:((Xaa^(neu)-Xaa⁺)_(x)(Xaa^(neu)-Xaa⁻)_(y))_(n)  (I)((Xaa^(neu)-Xaa⁻)_(x)(Xaa^(neu)-Xaa⁺)_(y))_(n)  (II)((Xaa⁺-Xaa^(neu))_(x)(Xaa⁻-X aa^(neu))_(y))_(n)  (III)((Xaa⁻-Xaa^(neu))_(x)(Xaa⁺-X aa^(neu))_(y))_(n)  (IV)Xaa^(neu) represents an amino acid residue having a neutral charge; Xaa⁺represents an amino acid residue having a positive charge; Xaa⁻represents an amino acid residue having a negative charge; x and y areintegers having a value of 1, 2, 3, or 4, independently; and n is aninteger having a value of 1-5. Peptides with modulus I (i.e., peptideshaving alternate positively and negatively charged R groups on one side(e.g., the polar face of the β-sheet) are described by each of FormulasI-IV, where x and y are 1. Peptides of modulus II (i.e., peptides havingtwo residues bearing one type of charge (e.g., a positive charge)followed by two residues bearing another type of charge (e.g., anegative charge)) are described by the same formulas where both x and yare 2. Examples of peptides of modulus III (i.e. peptides having threeresidues bearing one type of charge (e.g., a positive charge) followedby three residues bearing another type of charge (e.g., a negativecharge)) include, but are not limited to,

(SEQ ID NO: 57) RARARADADADA.

Other hydrophilic residues that form hydrogen bonds including, but notlimited to, asparagine and glutamine, may be incorporated into thepeptides. If the alanine residues in the peptides are changed to morehydrophobic residues, such as leucine, isoleucine, phenylalanine ortyrosine, the resulting peptides have a greater tendency toself-assemble and form peptide matrices with enhanced strength. Somepeptides that have similar amino acids compositions and lengths as thepeptides described here form alpha-helices and random-coils rather thanbeta-sheets and do not form macroscopic structures. Thus, in addition toself-complementarity, other factors are likely to be important for theformation of macroscopic structures, such as the peptide length, thedegree of intermolecular interaction, and the ability to form staggeredarrays.

Self-assembled structures can be formed that have varying degrees ofstiffness or elasticity. The structures typically have a low elasticmodulus (e.g., a modulus in the range of 1-10 kPa as measured bystandard methods, such as in a standard cone-plate rheometer). Lowvalues may be preferable, as they permit structure deformation as aresult of movement, in response to pressure, in the event of cellcontraction. More specifically, stiffness can be controlled in a varietyof ways, including by changing the length, sequence, and/orconcentration of the precursor molecules (e.g., self-assemblingpeptides). Other methods for increasing stiffness can also be employed.For example, one can attach, to the precursors, biotin molecules or anyother molecules that can be subsequently cross-linked or otherwisebonded to one another. The molecules (e.g., biotin) can be included atan N- or C-terminus of a peptide or attached to one or more residuesbetween the termini. Where biotin is used, cross-linking can be achievedby subsequent addition of avidin. Biotin-containing peptides or peptidescontaining other cross-linkable molecules are within the scope of thepresent invention. For example, amino acid residues with aromatic ringsmay be incorporated and cross-linked by exposure to UV light. The extentof crosslinking can be precisely controlled by applying the radiationfor a predetermined length of time to peptides of known sequence andconcentration. The extent of crosslinking can be determined by lightscattering, gel filtration, or scanning electron microscopy usingstandard methods. Furthermore, crosslinking can be examined by HPLC ormass spectrometry analysis of the structure after digestion with aprotease, such as matrix metalloproteases. Material strength may bedetermined before and after cross-linking. Regardless of whethercross-linking is achieved by a chemical agent or light energy, themolecules may be cross-linked in the course of creating a mold or whenpeptide-containing solutions are applied to the body.

The half-life (e.g., the in vivo half-life) of the structures can alsobe modulated by incorporating protease or peptidase cleavage sites intothe precursors that subsequently form a given structure. Proteases orpeptidases that occur naturally in vivo or that are introduced (e.g., bya surgeon) can then promote degradation by cleaving their cognatesubstrates.

Combinations of any of the modifications described here can be made. Forexample, self-assembling peptides that include a protease cleavage siteand a cysteine residue and/or a cross-linking agent, kits and devicescontaining them, and methods of using them can be utilized.

The peptide structures formed from any self-assembling peptides made byany process can be characterized using various biophysical and opticaltechniques, such as circular dichroism (CD), dynamic light scattering,Fourier transform infrared (FTIR), atomic force (tension) microscopy(ATM), scanning electron microscopy (SEM), and transmission electronmicroscopy (TEM). For example, biophysical methods can be used todetermine the degree of beta-sheet secondary structure in the peptidestructure. Filament and pore size, fiber diameter, length, elasticity,and volume fraction can be determined using quantitative image analysisof scanning and/or transmission electron micrographs. The structures canalso be examined using several standard mechanical testing techniques tomeasure the extent of swelling, the effect of pH and ion concentrationon structure formation, the level of hydration under various conditions,the tensile strength, as well as the manner in which variouscharacteristics change over the period of time required for thestructures to form and degrade. These methods allow one of ordinaryskill in the art to determine which of the various alternatives andpeptides described herein are most suitable for use in the variousmethods, and allow optimization of the various processes.

In another embodiment, the self-assembling materials can anchor orinteract with the structural extracellular matrix (ECM) at the edges ofblood vessels and/or tissues are described herein. These self-assemblingmaterials typically have hydrophobic and/or hydrophilic sections whichallow the material to react or interact with the glycoproteins found inthe ECM.

Preferably, the self-assembling materials when they break down, do notcause any secondary toxicity. Further, the break down product of theself-assembling materials would be suitable for the growth and repair ofthe surrounding tissues.

Peptidomimetics

Another class of materials that can self assemble are peptidomimetics.Peptidomimetics, as used herein, refers to molecules which mimic peptidestructure. Peptidomimetics have general features analogous to theirparent structures, polypeptides, such as amphiphilicity. Examples ofsuch peptidomimetic materials are described in Moore et al., Chem. Rev.101(12), 3893-4012 (2001).

The peptidomimetic materials can be classified into four categories:α-peptides, β-peptides, γ-peptides, and δ-peptides. Copolymers of thesepeptides can also be used.

Examples of α-peptide peptidomimetics include, but are not limited to,N,N′-linked oligoureas, oligopyrrolinones, oxazolidin-2-ones, azatidesand azapeptides.

Examples of β-peptides include, but are not limited to, β-peptidefoldamers, α-aminoxy acids, sulfur-containing β-peptide analogues, andhydrazino peptides.

Examples of γ-peptides include, but are not limited to, γ-peptidefoldamers, oligoureas, oligocarbamates, and phosphodiesters.

Examples of δ-peptides include, but are not limited to, alkene-basedδ-amino acids and carbopeptoids, such as pyranose-based carbopeptoidsand furanose-based carbopeptoids.

Backbones which can Adopt Helical or Sheet Conformations

Another class of compounds that self assemble includes oligomers havingbackbones which can adopt helical or sheet conformations. Example ofsuch compounds include, but are not limited to, compounds havingbackbones utilizing bipyridine segments, compounds having backbonesutilizing solvophobic interactions, compounds having backbones utilizingside chain interactions, compounds having backbones utilizing hydrogenbonding interactions, and compounds having backbones utilizing metalcoordination.

Examples of compounds containing backbones utilizing bipyridine segmentsinclude, but are not limited to, oligo(pyridine-pyrimidines),oligo(pyridine-pyrimidines) with hydrazal linkers, andpyridine-pyridazines.

Examples of compounds containing backbones utilizing solvophobicinteractions include, but are not limited to, oligoguanidines, aedamers(structures which take advantage of the stacking properties of aromaticelectron donor-acceptor interactions of covalently linked subunits) suchas oligomers containing 1,4,5,8-naphthalene-tetracarboxylic diimiderings and 1,5-dialkoxynaphthalene rings, and cyclophanes such assubstituted N-benzyl phenylpyridinium cyclophanes.

Examples of compounds containing backbones utilizing side chaininteractions include, but are not limited to, oligothiophenes such asolihothiophenes with chiral p-phenyl-oxazoline side chains, andoligo(m-phenylene-ethynylene)s.

Examples of compound containing backbones utilizing hydrogen bondinginteractions include, but are not limited to, aromatic amide backbonessuch as oligo(acylated 2,2′-bipyridine-3,3′-diamine)s andoligo(2,5-bis[2-aminophenyl]pyrazine)s, diaminopyridine backbonestemplated by cyanurate, and phenylene-pyridine-pyrimidine ethynylenebackbones templated by isophthalic acid.

Examples of compounds containing backbones utilizing metal coordinationinclude, but are not limited to, zinc bilinones, oligopyridinescomplexed with Co(II), Co(III), Cu(II), Ni(II), Pd(II), Cr(III), orY(III), oligo(m-phenylene ethynylene)s containing metal-coordinatingcyano groups, and hexapyrrins.

Nucleotidomimetics

Another class of molecules which can self assemble arenucleotidomimetics such as isomeric oligonucleotides, modifiedcarbohydrates, nucleotides with modified nucleotide linkages, andnucleotides with alternative nucleobases.

Examples of isomeric nucleotides include, but are not limited to,iso-RNA and iso-DNA and α-DNA (change in the anomeric configuration fromβ to α), alt-DNA, and 1-DNA.

Examples of modified carbohydrates include, but are not limited to,backbones with C 1′-bases connectivities such as tetrafuranosyloligonucleotides, pentopyranosyl oligonucleotides, and hexopyranosyloligonucleotides; backbones with C2′-base connectivities such asisonucleotides (repositioning of the base sugar connection from C1 tothe C2 position), HNAs (insertion of an additional methylene groupbetween the 04′ and C1′ position of a furanose), ANAs (incorporation ofa C3′-(S)-hydroxyl group), MNAs (inversion of the C3′-OH configurationfrom (S) in ANAs to (R)), CNAs (replacement of the O of the hexose witha methylene group), CeNAs (introduction of a 5′-6′ alkene within theanalogous ring), as well as other ring systems, torsionally restrictedoligonucleotides such as bicyclic oligonucleotides, LNAs (restriction ofthe pentofuranose backbone to the 3′-endo configuration), torsionallyflexible oligonucleotides such as base sugar extensions (insertion ofmethylene and ethylene groups into both α- and β-deoxynucleotides) andacyclic backbones (glycerol derivatives incorporating phosphodiesterlinkages).

Examples of nucleotides with modified nucleotide linkages include, butare not limited to, PNAs (peptide nucleic acids), NDPs(nucleo-δ-peptides), fused sugar-base backbones, and cationic linkages.

Examples of alternative nucleobases include, but are not limited to,nucleotides with alternative aromatic nucleobases.

Other Materials

Other materials which can self assemble include N-alkylacrylamideoligomers and di- and triblock co-polymers. N-alkylacrylamides canassume self-assembled into sheet-like structures (see Kendhale et al.,Chem Comm.). Examples of block copolymers include copolypeptides,polypeptide-PEGS, PEO-polybutadienes, PEG-polysaccharides, etc.

Another class of materials which are known to self-assemble aredendrimers. “Dendrimers”, as used herein, refers to branched polymerswith successive shells of branch units surrounding central core.Dendrimers can self-assemble through a variety of different mechanisms,such as hydrogen bonding, ionic interactions, hydrophobic interactions,solvent interaction, side chain interactions, and the like. Non-limitingexamples of self-assembling dendrimers are described in Zimmerman etal., Science, Vol. 271, No. 5252, 1095-1098 (1996); Zimmerman et al., J.Am. Chem. Soc., 124(46), 13757-13769 (2002); and Frechet, Proc. Nat.Acad. Sci., Vol. 99, No. 8, 4782-4787 (2002).

B. Formation of Self-Assembling Materials

Prior to self-assembly, the materials may be contained in (e.g.,dissolved in) a solution that is substantially free of ions (e.g.,monovalent ions) or that contains a sufficiently low concentration ofions to prevent significant self-assembly (e.g., a concentration of ionsless than 10, 5, 1, or 0.1 mM). Self-assembly may be initiated orenhanced at any subsequent time by the addition of an ionic solute ordiluent to a solution of the material or by a change in pH. For example,NaCl at a concentration of between approximately 5 mM and 5 M can inducethe assembly of macroscopic structures within a short period of time(e.g., within a few minutes). Lower concentrations of NaCl may alsoinduce assembly but at a slower rate. Alternatively, self-assembly maybe initiated or enhanced by introducing the materials (whether dry, in asemi-solid gel, or dissolved in a liquid solution that is substantiallyfree of ions) into a fluid (e.g., a physiological fluid such as blood orgastric juice) or an area (e.g., a body cavity such as the nose or mouthor a cavity exposed by a surgical procedure) comprising such ions. Thegel does not have to be preformed prior to application to the desiredsite. Generally, self-assembly is expected to occur upon contacting thematerials with such a solution in any manner.

A wide variety of ions, including anions and cations (whether divalent,monovalent, or trivalent), can be used. For example, one can promote aphase transition by exposure to monovalent cations such as Li⁺, Na⁺, K⁺,and Cs⁺. The concentration of such ions required to induce or enhanceself-assembly is typically at least 5 mM (e.g., at least 10, 20, or 50mM). Lower concentrations also facilitate assembly, although at areduced rate. When desired, self-assembling materials can be deliveredwith a hydrophobic material (e.g. a pharmaceutically acceptable oil) ina concentration that permits self-assembly, but at a reduced rate. Whenself-assembling materials are mixed with a hydrophobic agent such as anoil or lipid the assembly of the material forms different structures.The structures will appear like ice on a layer of oil. In some caseswhen another material is added, the material will assemble into variousother three dimensional structures that may be suitable for loading of atherapeutic agent. The hydrophilic part of the molecule will assemble insuch a way as to minimize hydrophobic-hydrophilic interaction, therebycreating a barrier between the two environments. Several experimentshave shown that the self-assembling materials will align on the surfaceof the oil like ice on water with the hydrophobic part of the moleculetoward the surface and the hydrophilic portion of the molecule facingaway from the oil, or will form toroidal like structures with thehydrophobic material contained inside. This type of behavior enables theencapsulation of therapeutics or other molecule of interested fordelivery in the body.

Alternatively, some of the materials described herein do not requireions to self-assemble but may self assemble due to interactions with asolvent, hydrophobic interactions, side chain interactions, hydrogenbonding, and the like.

Depending on the formulation and desired properties of the macroscopicstructure (e.g., the stiffness of the scaffold or the rate of itsformation), the concentration of precursors (e.g., self-assemblingmaterials) can vary from approximately 0.01% w/v (0.1 mg/ml) toapproximately 99.99% w/v (999.9 mg/ml), inclusive. For example, theconcentration prior to scaffold formation can be between approximately0.1% (1 mg/ml) and 10% (100 mg/ml), inclusive (e.g., about 0.1%-5%;0.5%-5%; 1.0%; 1.5%; 2.0%; 2.5%; 3.0%; or 4.0% or more). The precursors(e.g., self-assembling materials) can be formulated as powders andadministered in a powder form or resuspended. If dry, the materials canthen self-assemble following contact with bodily fluids (e.g., at a siteof injury).

The materials can be formed within regularly or irregularly-shapedmolds, which may include a body cavity or a portion of the body (e.g.,the lumen of a blood vessel) or which may be an inert material such asplastic or glass. The structures or scaffolds can be made to conform toa predetermined shape or to have a predetermined volume. To form astructure with a predetermined shape or volume (e.g., a desired geometryor dimension, including thin sheets or films), an aqueous solution ofthe material is placed in a pre-shaped casting mold, and the materialsare induced to self-assemble by the addition of a plurality of ions.Alternately, the ions may be added to the solution shortly beforeplacing the solution into the mold, provided that care is taken to placethe solution into the mold before substantial assembly occurs. Where themold is a tissue (e.g., the lumen of a blood vessel or othercompartment, whether in situ or not), the addition of an ionic solutionmay not be necessary. The resulting material characteristics, the timerequired for assembly, and the dimensions of the macroscopic structurethat forms are governed by the concentration and amount of solution thatis applied, the concentration of ions used to induce assembly of thestructure, and the dimensions of the casting apparatus. The scaffold canachieve a gel-like or substantially solid form at room temperature, andheat may be applied to facilitate the molding (e.g., one can heat asolution used in the molding process (e.g., a precursor-containingsolution) to a temperature ranging up to about body temperature(approximately 378 C.)). Once the scaffold has reached the desireddegree of firmness, it can be removed from the mold and used for apurpose described herein. Alternatively, the materials described hereinmay be used to anchor host tissue to a tissue matrix or scaffold. Forexample, the materials described herein can be used as a “glue” toanchor host tissue that is to be regenerated to a tissue matrix orscaffold to ensure that the matrix or scaffold stays in place in thelocal environment to which it is injected or implanted. Tissue matricesand scaffolds are well known in the art and can be prepared fromsynthetic, semi-synthetic, and/or natural materials.

Materials that assemble and/or undergo a phase transition (e.g., atransition from a liquid state to a semi-solid, gel, etc.) when theycome in contact with the body are useful in preventing the movement ofbodily substances. Self-assembly or phase transition is triggered bycomponents found in a subject's body (e.g., ions) or by physiological pHand is assisted by physiological temperatures. Self-assembly or phasetransition can begin when the compositions are exposed to or broughtinto contact with a subject's body and may be facilitated by the localapplication of heat to the area where the composition has been (or willbe) deposited. Based on studies to date, self-assembly occurs rapidlyupon contact with internal bodily tissues without the application ofadditional heat. The time required for effective assembly and/or phasetransition can occur in 60 seconds or less following contact with asubject's internal tissues or to conditions similar to those foundwithin the body (e.g., in 50, 40, 30, 20, or 10 seconds or less). Insome circumstances, such as where the concentration of self-assemblingagents in the composition is low or where the movement of the bodilysubstance is substantial, self-assembly or phase transition may takelonger to achieve the desired effect, for example, up to a minute, 5minutes, 10 minutes, 30 minutes, an hour, or longer. For example, asolution containing a self-assembling peptide applied to sites of bloodvessel transection in the brain, liver, or muscle provided completehemostasis within times as short as 10 seconds following application.Ion-containing solutions may be preferred when the compositions are usedto protect a subject from contamination, as phase transitions do notoccur, or do not readily occur, when non-ionic compositions contactintact skin.

The compositions can form structures that are substantially rigid (e.g.,solid or nearly solid) or that assume a definite shape and volume (e.g.,structures that conform to the shape and volume of the location to whicha liquid composition was administered, whether in vivo or ex vivo). Thesolidified material may be somewhat deformable or compressible afterassembly or phase transition, but will not substantially flow from onearea to another, as compositions at a different point along the liquidto solid continuum may do, which may be due, at least in part, to theirability to undergo phase transitions. As a result, the compositions canbe used to prevent the movement of a bodily substance in a subject inneed thereof. Self-assembly can also be achieved ex vivo by exposure toconditions within a certain range of physiological values (e.g.,conditions appropriate for cell or tissue culture). While liquidformulations are readily dispensed, the compositions administered mayalso be in a gel form that may become stiffer upon contact with thesubject's body.

The concentration of the self-assembling materials in any givenformulation can vary and can be between approximately 0.1% (1 mg/ml) and10% (100 mg/ml), inclusive. For example, the concentration of theself-assembling peptides (e.g., in a liquid formulation) can beapproximately 0.1-3.0% (1-30 mg/ml) (e.g., 0.1-1.0%; 1.0-2.0%; 2.0-3.0%or 1.0-3.0%). The concentration of self-assembling materials can behigher in stock solutions and in solid (e.g., powdered) formulations. Insolid preparations, the concentration of self-assembling materials canapproach 100% (e.g., the concentration of self-assembling peptides canbe 95, 96, 97, 98, 99% or more (e.g., 99.99%) of the composition).Whether in liquid or solid form, the materials can be brought to thedesired concentration prior to use by addition of a diluent (e.g.,deionized water), powder, wetting agent, or a therapeutic, diagnostic orprophylactic agent.

Regardless of the precise nature of the self-assembling materials, uponexposure to conditions such as those described herein, the materials canform membranous two- or three-dimensional structures including a stablemacroscopic porous matrix having ordered interwoven nanofibers (e.g.,fibers approximately 10-20 nm in diameter, with a pore size of about50-100 nm in a linear dimension). Three-dimensional macroscopic matricescan have dimensions large enough to be visible under low magnification(e.g., about 10-fold or less), and the membranous structures can bevisible to the naked eye, even if transparent. Althoughthree-dimensional, the structures can be exceedingly thin, including alimited number of layers of molecules (e.g., 2, 3, or more layers ofmolecules). Typically, each dimension of a given structure will be atleast 10 μm in size (e.g., two dimensions of at least 100-1000 μm insize (e.g., 1-10 mm, 10-100 mm, or more)). The relevant dimensions maybe expressed as length, width, depth, breadth, height, radius, diameter,or circumference in the case of structures that have a substantiallyregular shape (e.g., where the structure is a sphere, cylinder, cube, orthe like) or an approximation of any of the foregoing where thestructures do not have a regular shape.

The self-assembling materials can form a hydrated material whencontacted with water under conditions such as those described herein(e.g., in the presence of a sufficient concentration (e.g.,physiological concentrations) of ions (e.g., monovalent cations)). Thematerials may have a high water content (e.g., approximately 95% or more(e.g., approximately 97%, 98%, 99% or more)), and the compositions canbe hydrated but not substantially self-assembled. A given value may be“approximate” in recognition of the fact that measurements can varydepending, for example, on the circumstances under which they are madeand the skill of the person taking the measurement. Generally, a firstvalue is approximately equal to a second when the first falls within 10%of the second (whether greater than or less than) unless it is otherwiseclear from the context that a value is not approximate or where, forexample, such value would exceed 100% of a possible value.

The properties and mechanical strength of the structures or scaffoldscan be controlled as required through manipulation of the componentstherein. For example, the stiffness of an assembled gel can be increasedby increasing the concentration of self-assembling materials therein.The sequences, characteristics, and properties of the materials and thestructures formed by them upon self-assembly are discussed furtherbelow.

The compositions can be formulated as concentrated stocks or in dryform, and these can be diluted or dissolved to form biocompatiblecompositions, which are substantially non-toxic to biological cells invitro or in vivo. For example, the compositions can contain materials inquantities that do not elicit a significant deleterious effect on therecipient's body (e.g., a prohibitively severe immunological orinflammatory reaction, or unacceptable scar tissue formation).

When a solution containing non-assembled materials is laid down on abiological tissue, the materials having sufficient proximity to thetissue assemble, causing the solution to gel. Any solution that remainsdistant from the tissue remains liquid, as the self-assembling materialshave not yet been exposed to conditions that promote their assembly. Asthe material is disturbed (e.g., by performing a surgical procedure),liquid material appears to gel as it comes into sufficient contact withthe body. At times, the compositions can take on characteristics rangingfrom a liquid to those of a solid, appearing gel- or salve-like or as aslurry).

The self assembling material may further contain a tissue specificcomponent. The tissue specific component can be peptides,polysaccharides, or glycoproteins that are specific for eye, brain, orskin cells. For example, cell surface carbohydrates are major componentsof the outer surface of mammalian cells and are very oftencharacteristic of cell types. It is assumed that cell type-specificcarbohydrates are involved in cell-cell interaction. The tissue specificcomponent can therefore, target these cell specific surfacecarbohydrates.

(SEQ ID NO: 58)   1 G G G G G D G D G D G D G D G D (SEQ ID NO: 59)  2 G G G G G E G E G E G E G E G E (SEQ ID NO: 60)  3 G G G G G K G K G K G K G K G K (SEQ ID NO: 61)  4 G G G G G R G R G R G R G R G R (SEQ ID NO: 62)  5 G G G G G H G H G H G H G H G H (SEQ ID NO: 63)  6 A A A A A D A D A D A D A D A D (SEQ ID NO: 64)  7 A A A A A E A E A E A E A E A E (SEQ ID NO: 65)  8 A A A A A K A K A K A K A K A K (SEQ ID NO: 66)  9 A A A A A R A R A R A R A R A R (SEQ ID NO: 67) 10 A A A A A H A H A H A H A H A H (SEQ ID NO: 68) 11 V V V V V D V D V D V D V D V D (SEQ ID NO: 69) 12 V V V V V E V E V E V E V E V E (SEQ ID NO: 70) 13 V V V V V K V K V K V K V K V K (SEQ ID NO: 71) 14 V V V V V R V R V R V R V R V R (SEQ ID NO: 72) 15 V V V V V H V H V H V H V H V H (SEQ ID NO: 73) 16 L L L L L D L D L D L D L D L D (SEQ ID NO: 74) 17 L L L L L E L E L E L E L E L E (SEQ ID NO: 75) 18 L L L L L K L K L K L K L K L K (SEQ ID NO: 76) 19 L L L L L R L R L R L R L R L R (SEQ ID NO: 77) 20 L L L L L H L H L H L H L H L H (SEQ ID NO: 78) 21 I I I I I D I D I D I D I D I D (SEQ ID NO: 79) 22 I I I I I E I E I E I E I E I E (SEQ ID NO: 80) 23 I I I I I K I K I K I K I K I K (SEQ ID NO: 81) 24 I I I I I R I R I R I R I R I R (SEQ ID NO: 82) 25 I I I I I H I H I H I H I H I H (SEQ ID NO: 83) 26 M M M M M D M D M D M D M D M D (SEQ ID NO: 84) 27 M M M M M E M E M E M E M E M E (SEQ ID NO: 85) 28 M M M M M K M K M K M K M K M K (SEQ ID NO: 86) 29 M M M M M R M R M R M R M R M R (SEQ ID NO: 87) 30 M M M M M H M H M H M H M H M H (SEQ ID NO: 88) 31 F F F F F D F D F D F D F D F D (SEQ ID NO: 89) 32 F F F F F E F E F E F E F E F E (SEQ ID NO: 90) 33 F F F F F K F K F K F K F K F K (SEQ ID NO: 91) 34 F F F F F R F R F R F R F R F R (SEQ ID NO: 92) 35 F F F F F H F H F H F H F H F H (SEQ ID NO: 93) 36 W W W W W D W D W D W D W D W D (SEQ ID NO: 94) 37 W W W W W E W E W E W E W E W E (SEQ ID NO: 95) 38 W W W W W K W K W K W K W K W K (SEQ ID NO: 96) 39 W W W W W R W R W R W R W R W R (SEQ ID NO: 97) 40 W W W W W H W H W H W H W H W H (SEQ ID NO: 98) 41 P P P P P D P D P D P D P D P D (SEQ ID NO: 98) 42 P P P P P E P E P E P E P E P E (SEQ ID NO: 100) 43 P P P P P K P K P K P K P K P K (SEQ ID NO: 101) 44 P P P P P R P R P R P R P R P R (SEQ ID NO: 102) 45 P P P P P H P H P H P H P H P H (SEQ ID NO: 103) 46 A A A A A R A D A R A D A R A D (SEQ ID NO: 104) 47 A A A A A R A R A D A D A R A R (SEQ ID NO: 105) 48 A A A A A E A K A E A K A E A K (SEQ ID NO: 106) 49 A A A A A E A E A K A K A E A E (SEQ ID NO: 107) 50 A A A A A R A E A R A E A R A E (SEQ ID NO: 108) 51 A A A A A R A R A E A E A R A E (SEQ ID NO: 109) 52 A A A A A K A D A K A D A K A D (SEQ ID NO: 110) 53 A A A A A E A H A E A H A E A H (SEQ ID NO: 111) 54 A A A A A E A E A H A H A E A E (SEQ ID NO: 112) 55 A A A A A R A R A R A R A R A R (SEQ ID NO: 113) 56 A A A A A R A R A R A R A D A D (SEQ ID NO: 114) 57 A A A A A R A R A R A D A D A D (SEQ ID NO: 115) 58 A A A A A H A D A N A D A H A D (SEQ ID NO: 116) 59 A A A A A H A H A H A H A H A H (SEQ ID NO: 117) 60 A A A A A H A D A D A H A D A D (SEQ ID NO: 118) 61 A A A A A H A E A E A H A E A E (SEQ ID NO: 119) 62 G G G G G R G D G R G D G R G D (SEQ ID NO: 120) 63 G G G G G R G R G D G D G R G R (SEQ ID NO: 121) 64 G G G G G E G K G E G K G E G K (SEQ ID NO: 122) 65 G G G G G E G E G K G K G E G E (SEQ ID NO: 123) 66 G G G G G R G E G R G E G R G E (SEQ ID NO: 124) 67 G G G G G R G R G E G E G R G E (SEQ ID NO: 125) 68 G G G G G K G D G K G D G K G D (SEQ ID NO: 126) 69 G G G G G E G H G E G H G E G H (SEQ ID NO: 127) 70 G G G G G E G E G H G H G E G E (SEQ ID NO: 128) 71 G G G G G R G R G R G R G R G R (SEQ ID NO: 129) 72 G G G G G R G R G R G R G D G D (SEQ ID NO: 130) 73 G G G G G R G R G R G D G D G D (SEQ ID NO: 131) 74 G G G G G H G D G H G D G H G D (SEQ ID NO: 132) 75 G G G G G H G H G H G H G H G H (SEQ ID NO: 133) 76 G G G G G H G D G D G H G D G D (SEQ ID NO: 134) 77 G G G G G H G E G E G H G E G E (SEQ ID NO: 135) 78 V V V V V R V D V R V D V R V D (SEQ ID NO: 136) 79 V V V V V R V R V D V D V R V R (SEQ ID NO: 137) 80 V V V V V E V K V E V K V E V K (SEQ ID NO: 138) 81 V V V V V E V E V K V K V E V E (SEQ ID NO: 139) 82 V V V V V R V E V R V E V R V E (SEQ ID NO: 140) 83 V V V V V R V R V E V E V R V E (SEQ ID NO: 141) 84 V V V V V K V D V K V D V K V D (SEQ ID NO: 142) 85 V V V V V E V H V E V H V E V H (SEQ ID NO: 143) 86 V V V V V E V E V H V H V E V E (SEQ ID NO: 144) 87 V V V V V R V R V R V R V R V R (SEQ ID NO: 145) 88 V V V V V R V R V R V R V D V D (SEQ ID NO: 146) 89 V V V V V R V R V R V D V D V D (SEQ ID NO: 147) 90 V V V V V H V D V H V D V H V D (SEQ ID NO: 148) 91 V V V V V H V H V H V H V H V H (SEQ ID NO: 149) 92 V V V V V H V D V D V H V D V D (SEQ ID NO: 150) 93 V V V V V H V E V E V H V E V E (SEQ ID NO: 151) 94 L L L L L R L D L R L D L R L D (SEQ ID NO: 152) 95 L L L L L R L R L D L D L R L R (SEQ ID NO: 153) 96 L L L L L E L K L E L K L E L K (SEQ ID NO: 154) 97 L L L L L E L E L K L K L E L E (SEQ ID NO: 155) 98 L L L L L R L E L R L E L R L E (SEQ ID NO: 156) 99 L L L L L R L R L E L E L R L E (SEQ ID NO: 157)100 L L L L L K L D L K L D L K L D (SEQ ID NO: 158)101 L L L L L E L H L E L H L E L H (SEQ ID NO: 159)102 L L L L L E L E L H L H L E L E (SEQ ID NO: 160)103 L L L L L R L R L R L R L R L R (SEQ ID NO: 161)104 L L L L L R L R L R L R L D L D (SEQ ID NO: 162)105 L L L L L R L R L R L D L D L D (SEQ ID NO: 163)106 L L L L L H L D L H L D L H L D (SEQ ID NO: 164)107 L L L L L H L H L H L H L H L H (SEQ ID NO: 165)108 L L L L L H L D L D L H L D L D (SEQ ID NO: 166)109 L L L L L H L E L E L H L E L E (SEQ ID NO: 167)110 I I I I I R I D I R I D I R I D (SEQ ID NO: 168)111 I I I I I R I R I D I D I R I R (SEQ ID NO: 169)112 I I I I I E I K I E I K I E I K (SEQ ID NO: 170)113 I I I I I E I E I K I K I E I E (SEQ ID NO: 171)114 I I I I I R I E I R I E I R I E (SEQ ID NO: 172)115 I I I I I R I R I E I E I R I E (SEQ ID NO: 173)116 I I I I I K I D I K I D I K I D (SEQ ID NO: 174)117 I I I I I E I H I E I H I E I H (SEQ ID NO: 175)118 I I I I I E I E I H I H I E I E (SEQ ID NO: 176)119 I I I I I R I R I R I R I R I R (SEQ ID NO: 177)120 I I I I I R I R I R I R I D I D (SEQ ID NO: 178)121 I I I I I R I R I R I D I D I D (SEQ ID NO: 179)122 I I I I I H I D I H I D I H I D (SEQ ID NO: 180)123 I I I I I H I H I H I H I H I H (SEQ ID NO: 181)124 I I I I I H I D I D I H I D I D (SEQ ID NO: 182)125 I I I I I H I E I E I H I E I E (SEQ ID NO: 183)126 M M M M M R M D M R M D M R M D (SEQ ID NO: 184)127 M M M M M R M R M D M D M R M R (SEQ ID NO: 185)128 M M M M M E M K M E M K M E M K (SEQ ID NO: 186)129 M M M M M E M E M K M K M E M E (SEQ ID NO: 187)130 M M M M M R M E M R M E M R M E (SEQ ID NO: 188)131 M M M M M R M R M E M E M R M E (SEQ ID NO: 189)132 M M M M M K M D M K M D M K M D (SEQ ID NO: 190)133 M M M M M E M H M E M H M E M H (SEQ ID NO: 191)134 M M M M M E M E M H M H M E M E (SEQ ID NO: 192)135 M M M M M R M R M R M R M R M R (SEQ ID NO: 193)136 M M M M M R M R M R M R M D M D (SEQ ID NO: 194)137 M M M M M R M R M R M D M D M D (SEQ ID NO: 195)138 M M M M M H M D M H M D M H M D (SEQ ID NO: 196)139 M M M M M H M H M H M H M H M H (SEQ ID NO: 197)140 M M M M M H M D M D M H M D M D (SEQ ID NO: 198)141 M M M M M H M E M E M H M E M E (SEQ ID NO: 199)142 F F F F F R F D F R F D F R F D (SEQ ID NO: 200)143 F F F F F R F R F D F D F R F R (SEQ ID NO: 201)144 F F F F F E F K F E F K F E F K (SEQ ID NO: 202)145 F F F F F E F E F K F K F E F E (SEQ ID NO: 203)146 F F F F F R F E F R F E F R F E (SEQ ID NO: 204)147 F F F F F R F R F E F E F R F E (SEQ ID NO: 205)148 F F F F F K F D F K F D F K F D (SEQ ID NO: 206)149 F F F F F E F H F E F H F E F H (SEQ ID NO: 207)150 F F F F F E F E F H F H F E F E (SEQ ID NO: 208)151 F F F F F R F R F R F R F R F R (SEQ ID NO: 209)152 F F F F F R F R F R F R F D F D (SEQ ID NO: 210)153 F F F F F R F R F R F D F D F D (SEQ ID NO: 211)154 F F F F F H F D F H F D F H F D (SEQ ID NO: 212)155 F F F F F H F H F H F H F H F H (SEQ ID NO: 213)156 F F F F F H F D F D F H F D F D (SEQ ID NO: 214)157 F F F F F H F E F E F H F E F E (SEQ ID NO: 215)158 W W W W W R W D W R W D W R W D (SEQ ID NO: 216)159 W W W W W R W R W D W D W R W R (SEQ ID NO: 217)160 W W W W W E W K W E W K W E W K (SEQ ID NO: 218)161 W W W W W E W E W K W K W E W E (SEQ ID NO: 219)162 W W W W W R W E W R W E W R W E (SEQ ID NO: 220)163 W W W W W R W R W E W E W R W E (SEQ ID NO: 221)164 W W W W W K W D W K W D W K W D (SEQ ID NO: 222)165 W W W W W E W H W E W H W E W H (SEQ ID NO: 223)166 W W W W W E W E W H W H W E W E (SEQ ID NO: 224)167 W W W W W R W R W R W R W R W R (SEQ ID NO: 225)168 W W W W W R W R W R W R W D W D (SEQ ID NO: 226)169 W W W W W R W R W R W D W D W D (SEQ ID NO: 227)170 W W W W W H W D W H W D W H W D (SEQ ID NO: 228)171 W W W W W H W H W H W H W H W H (SEQ ID NO: 229)172 W W W W W H W D W D W H W D W D (SEQ ID NO: 230)173 W W W W W H W E W E W H W E W E (SEQ ID NO: 231)174 P P P P P R P D P R P D P R P D (SEQ ID NO: 232)175 P P P P P R P R P D P D P R P R (SEQ ID NO: 233)176 P P P P P E P K P E P K P E P K (SEQ ID NO: 234)177 P P P P P E P E P K P K P E P E (SEQ ID NO: 235)178 P P P P P R P E P R P E P R P E (SEQ ID NO: 236)179 P P P P P R P R P E P E P R P E (SEQ ID NO: 237)180 P P P P P K P D P K P D P K P D (SEQ ID NO: 238)181 P P P P P E P H P E P H P E P H (SEQ ID NO: 239)182 P P P P P E P E P H P H P E P E (SEQ ID NO: 240)183 P P P P P R P R P R P R P R P R (SEQ ID NO: 241)184 P P P P P R P R P R P R P D P D (SEQ ID NO: 242)185 P P P P P R P R P R P D P D P D (SEQ ID NO: 243)186 P P P P P H P D P H P D P H P D (SEQ ID NO: 244)187 P P P P P H P H P H P H P H P H (SEQ ID NO: 245)188 P P P P P H P D P D P H P D P D (SEQ ID NO: 246)189 P P P P P H P E P E P H P E P E (SEQ ID NO: 247)190 S S S S S R S D S R S D S R S D (SEQ ID NO: 248)191 S S S S S R S R S D S D S R S R (SEQ ID NO: 249)192 S S S S S E S K S E S K S E S K (SEQ ID NO: 250)193 S S S S S E S E S K S K S E S E (SEQ ID NO: 251)194 S S S S S R S E S R S E S R S E (SEQ ID NO: 252)195 S S S S S R S R S E S E S R S E (SEQ ID NO: 253)196 S S S S S K S D S K S D S K S D (SEQ ID NO: 254)197 S S S S S E S H S E S H S E S H (SEQ ID NO: 255)198 S S S S S E S E S H S H S E S E (SEQ ID NO: 256)199 S S S S S R S R S R S R S R S R (SEQ ID NO: 257)200 S S S S S R S R S R S R S D S D (SEQ ID NO: 258)201 S S S S S R S R S R S D S D S D (SEQ ID NO: 259)202 S S S S S H S D S H S D S H S D (SEQ ID NO: 260)203 S S S S S H S H S H S H S H S H (SEQ ID NO: 261)204 S S S S S H S D S D S H S D S D (SEQ ID NO: 262)205 S S S S S H S E S E S H S E S E (SEQ ID NO: 263)206 T T T T T R T D T R T D T R T D (SEQ ID NO: 264)207 T T T T T R T R T D T D T R T R (SEQ ID NO: 265)208 T T T T T E T K T E T K T E T K (SEQ ID NO: 266)209 T T T T T E T E T K T K T E T E (SEQ ID NO: 267)210 T T T T T R T E T R T E T R T E (SEQ ID NO: 268)211 T T T T T R T R T E T E T R T E (SEQ ID NO: 269)212 T T T T T K T D T K T D T K T D (SEQ ID NO: 270)213 T T T T T E T H T E T H T E T H (SEQ ID NO: 271)214 T T T T T E T E T H T H T E T E (SEQ ID NO: 272)215 T T T T T R T R T R T R T R T R (SEQ ID NO: 273)216 T T T T T R T R T R T R T D T D (SEQ ID NO: 274)217 T T T T T R T R T R T D T D T D (SEQ ID NO: 275)218 T T T T T H T D T H T D T H T D (SEQ ID NO: 276)219 T T T T T H T H T H T H T H T H (SEQ ID NO: 277)220 T T T T T H T D T D T H T D T D (SEQ ID NO: 278)221 T T T T T H T E T E T H T E T E (SEQ ID NO: 279)222 C C C C C R C D C R C D C R C D (SEQ ID NO: 280)223 C C C C C R C R C D C D C R C R (SEQ ID NO: 281)224 C C C C C E C K C E C K C E C K (SEQ ID NO: 282)225 C C C C C E C E C K C K C E C E (SEQ ID NO: 283)226 C C C C C R C E C R C E C R C E (SEQ ID NO: 284)227 C C C C C R C R C E C E C R C E (SEQ ID NO: 285)228 C C C C C K C D C K C D C K C D (SEQ ID NO: 286)229 C C C C C E C H C E C H C E C H (SEQ ID NO: 287)230 C C C C C E C E C H C H C E C E (SEQ ID NO: 288)231 C C C C C R C R C R C R C R C R (SEQ ID NO: 289)232 C C C C C R C R C R C R C D C D (SEQ ID NO: 290)233 C C C C C R C R C R C D C D C D (SEQ ID NO: 291)234 C C C C C H C D C H C D C H C D (SEQ ID NO: 292)235 C C C C C H C H C H C H C H C H (SEQ ID NO: 293)236 C C C C C H C D C D C H C D C D (SEQ ID NO: 294)237 C C C C C H C E C E C H C E C E (SEQ ID NO: 295)238 Y Y Y Y Y R Y D Y R Y D Y R Y D (SEQ ID NO: 296)239 Y Y Y Y Y R Y R Y D Y D Y R Y R (SEQ ID NO: 297)240 Y Y YY Y E Y K Y E Y K Y E Y K (SEQ ID NO: 298)241 Y Y Y Y Y E Y E Y K Y K Y E Y E (SEQ ID NO: 299)242 Y Y Y Y Y R Y E Y R Y E Y R Y E (SEQ ID NO: 300)243 Y Y Y Y Y R Y R Y E Y E Y R Y E (SEQ ID NO: 301)244 Y Y Y Y Y K Y D Y K Y D Y K Y D (SEQ ID NO: 302)245 Y Y Y Y Y E Y H Y E Y H Y E Y H (SEQ ID NO: 303)246 Y Y Y Y Y E Y E Y H Y H Y E Y E (SEQ ID NO: 304)247 Y Y Y Y Y R Y R Y R Y R Y R Y R (SEQ ID NO: 305)248 Y Y Y Y Y R Y R Y R Y R Y D Y D (SEQ ID NO: 306)249 Y Y Y Y Y R Y R Y R Y D Y D Y D (SEQ ID NO: 307)250 Y Y Y Y Y H Y D Y H Y D Y H Y D (SEQ ID NO: 308)251 Y Y Y Y Y H Y H Y H Y H Y H Y H (SEQ ID NO: 309)252 Y Y Y Y Y H Y D Y D Y H Y D Y D (SEQ ID NO: 310)253 Y Y Y Y Y H Y E Y E Y H Y E Y E (SEQ ID NO: 311)254 N N N N N R N D N R N D N R N D (SEQ ID NO: 312)255 N N N N N R N R N D N D N R N R (SEQ ID NO: 313)256 N N N N N E N K N E N K N E N K (SEQ ID NO: 314)257 N N N N N E N E N K N K N E N E (SEQ ID NO: 315)258 N N N N N R N E N R N E N R N E (SEQ ID NO: 316)259 N N N N N R N R N E N E N R N E (SEQ ID NO: 317)260 N N N N N K N D N K N D N K N D (SEQ ID NO: 318)261 N N N N N E N H N E N H N E N H (SEQ ID NO: 319)262 N N N N N E N E N H N H N E N E (SEQ ID NO: 320)263 N N N N N R N R N R N R N R N R (SEQ ID NO: 321)264 N N N N N R N R N R N R N D N D (SEQ ID NO: 322)265 N N N N N R N R N R N D N D N D (SEQ ID NO: 323)266 N N N N N H N D N H N D N H N D (SEQ ID NO: 324)267 N N N N N H N H N H N H N H N H (SEQ ID NO: 325)268 N N N N N H N D N D N H N D N D (SEQ ID NO: 326)269 N N N N N H N E N E N H N E N E (SEQ ID NO: 327)270 Q Q Q Q Q R Q D Q R Q D Q R Q D (SEQ ID NO: 328)271 Q Q Q Q Q R Q R Q D Q D Q R Q R (SEQ ID NO: 329)272 Q Q Q Q Q E C K Q E Q K Q E Q K (SEQ ID NO: 330)273 Q Q Q Q Q E Q E Q K Q K Q E Q E (SEQ ID NO: 331)274 Q Q Q Q Q R Q E Q R Q E Q R Q E (SEQ ID NO: 332)275 Q Q Q Q Q R Q R Q E Q E Q R Q E (SEQ ID NO: 333)276 Q Q Q Q Q K Q D Q K Q D Q K Q D (SEQ ID NO: 334)277 Q Q Q Q Q E Q H Q E Q H Q E Q H (SEQ ID NO: 335)278 Q Q Q Q Q E Q E Q H Q H Q E Q E (SEQ ID NO: 336)279 Q Q Q Q Q R Q R Q R Q R Q R Q R (SEQ ID NO: 337)280 Q Q Q Q Q R Q R Q R Q R Q D Q D (SEQ ID NO: 338)281 Q Q Q Q Q R Q R Q R C D Q D Q D (SEQ ID NO: 339)282 Q Q Q Q Q H Q D Q H Q D Q H Q D (SEQ ID NO: 340)283 Q Q Q Q Q H Q H Q H Q H Q H Q H (SEQ ID NO: 341)284 Q Q Q Q Q H Q D Q D Q H Q D Q D (SEQ ID NO: 342)285 Q Q Q Q Q H Q E Q E Q H Q E Q E

B. Therapeutic, Prophylactic and Diagnostic Agents

The formulations may also include other therapeutic, prophylactic ordiagnostic agents. In a preferred embodiment, these may beanti-inflammatories, vasoactive agents, anti-infectives, anesthetics,growth factors, vitamin d, nutrients, and/or cells.

These can be peptides or proteins, polysaccharides or saccharides,nucleic acids nucleotides, proteoglycan, lipid, carbohydrate, or a smallmolecule, typically an organic compound, having multiple carbon-carbonbonds that may be isolated from nature or prepared via chemicalsynthesis. Small molecules have relatively low molecular weights (e.g.,less than about 1500 g/mol) and are not peptides or nucleic acids. Thesubstance can also be a biomolecule, which is a molecule such as apeptide, proteoglycan, lipid, carbohydrate, or nucleic acid havingcharacteristics typical of molecules found in living organisms Likesmall molecules, biomolecules can be naturally occurring or may beartificial (i.e., they may be molecules that have not been found innature). For example, a protein having a sequence that has not beenfound in nature (e.g., one that does not occur in a publicly availabledatabase of sequences) or that has a known sequence modified in anunnatural way by a human hand (e.g., a sequence modified by altering apost-translational process such as glycosylation) is an artificialbiomolecule. Nucleic acid molecules encoding such proteins (e.g., anoligonucleotide, optionally contained within an expression vector) arealso biomolecules and can be incorporated into the compositionsdescribed herein. For example, a composition can include a plurality ofself-assembling materials and cells that express, or that are engineeredto express, a protein biomolecule (by virtue of containing a nucleicacid sequence that encodes the protein biomolecule).

Many different therapeutic, prophylactic or diagnostic agents can beincorporated into the formulation. Representative vasoconstrictorsinclude epinephrine and phenylephrine; representative coloring agentsinclude arsenazo III, chlorophosphonazo III, antipyrylazo 111, murexide,Eriochrome Black T, Eriochrome Blue SE, oxyacetazo I, carboxyazo III,tropolone, methylthymol blue, and Mordant Black 32; representativeanesthetic agents include benzocaine, bupivacaine, butamben picrate,chloroprocaine, cocaine, curate, dibucaine, dyclonine, etidocaine,lidocaine, mepivacaine, pramoxine, prilocaine, procaine, propoxycaine,ropivacaine, tetracaine, or combinations thereof. Local application ofthe anesthetic agent may be all that is required in some situations, forexample, for a burn or other wound to the skin, including decubitusulcers; wounds, such as cancer sores; or for minimally invasivesurgeries. Combining local anesthetics with the self-assemblingmaterials, whether combined by virtue of being present in the sameformulation or by virtue of co-administration, can help contain theanesthetic within the body and reduce the amount entering thecirculation.

Vasoconstrictors such as phenylephrine can be included to prolong theeffect of local anesthesia (e.g., 0.1-0.5% phenylephrine). Analgesicagents other than a local anesthetic agent, such as steroids,non-steroidal anti-inflammatory agents like indomethacin, plateletactivating factor (PAF) inhibitors such as lexipafant, CV 3988, and/orPAF receptor inhibitors such as SRI 63-441. An anti-infective orantimicrobial agent (e.g., an antibiotic, antibacterial, antiviral, orantifungal agent) can be included for either systemic or localadministration. Examples include (β-lactam antibiotics such aspenicillins and cephalosporins and other inhibitors of cell wallsynthesis such as vancomycin, chloramphenicol, tetracyclines,macrolides, clindamyin, streptogramins, aminoglycosides, spectinomycin,sulfonamides, trimethoprim, quinolones, amphotericin B, flucytosine,azoles such as ketoconazole, itraconazole, fluconazole, clotrimazole,and miconazole, griseofulvin, terbinafine, and nystatin. Theantimicrobial can be topically administered (e.g., to treat skininfections or burns, or to help prevent infection at a site of catheterinsertion (e.g., an intravenous catheter), for example, kanamycin,neomycin, bacitracin, polymixin, topical sulfonamides such as mafenideacetate or silver sulfadiazine, or gentamicin sulfate. The antimicrobialcan also be a broad spectrum agent. For example, a second, third, orfourth generation cephalosporin can be used. These agents may be activeagainst a wide range of bacteria including both gram positive and gramnegative species. Such antibacterial agents may be particularlyappropriate where the present scaffolds are used to inhibit movement ofintestinal contents such as during intestinal resection or other surgerythat purposefully or accidentally disturbs the integrity of theintestinal wall. One of ordinary skill in the art will be able to selectappropriate antimicrobial agents by considering factors such as thepatient's history (e.g., any history of an allergic reaction to suchagents), the location to which the peptides are to be applied, the typeof infectious agent likely to be present, and so forth.

Any of the compositions described herein, whether they contain onlyself-assembling precursors or precursors and one or more bioactivemolecules (and whether in a liquid, semi-solid, or solid farm), caninclude a coloring agent. Suitable coloring agents include commerciallyavailable food colorings, natural and synthetic dyes, and fluorescentmolecules. Preferably, the coloring agent is nontoxic or is included atsuch low concentrations as to minimize any toxic effect. The use of acoloring agent allows for improved visualization of an area that iscovered by a structure or scaffold and can facilitate removal, if suchremoval is desired. The coloring agent can be one that changes colorwhen it comes into contact with a contaminated area (e.g., a colorchange may be triggered by the contamination itself (e.g., by the bloodor bacteria present at a wound site)). For example, a metabolic productof a bacterium may trigger a color change. Conditions such as pH orredox state induced by contaminants may also be detected. Exemplaryindicators include arsenzazo III, chlorophosphonazo III, antipyrylazoIII, murexide, Eriochrome Black T and Eriochrome Blue SE for Mg²⁺,oxyacetazo I, carboxyazo III, tropolone, methylthymol blue, and MordantBlack 32. AlamarBlue, a redox indicator, and phenol red are also of usein the compositions and methods.

Many other active agents can be included in the compositions. Forexample, a number of growth factors can be included to accelerate one ormore aspects of healing (e.g., angiogenesis, cell migration, processextension, and cell proliferation). These types of compositions can be“included” as others can, by virtue of inclusion in the compositions orby virtue of co-administration in the present methods. Examples includevascular endothelial growth factor (VEGF), a transforming growth factor(TGF) such as transforming growth factor p, a platelet derived growthfactor (PDGF), an epidermal growth factor (EGF), a nerve growth factor(NGF), an insulin-like growth factor (e.g., insulin-like growth factorI), a glial growth factor (GGF), a fibroblast growth factor (FGF), etc.It will be appreciated that in many cases these terms refer to a varietyof different molecular species. For example, several transforming growthfactor R species are known in the art. One of ordinary skill in the artwill be guided in the selection of an appropriate growth factor byconsidering, for example, the site at which the composition is to beadministered. For example, an EGF can be included in compositionsapplied to the skin; an NGF and/or GGF can be included in compositionsapplied to nerves or the nervous system; and so forth.

The growth factor or another agent can be a chemotactic substance, whichhas the ability, in vivo or in cell culture, to recruit cells to a siteat which the substance is present. The cells recruited may have thepotential to contribute to the formation of new tissue or to repairexisting, damaged tissue (e.g., by contributing structurally and/orfunctionally to the tissue (e.g., by providing growth factors orcontributing to a desirable immune response)). Certain chemotacticsubstances can also function as proliferation agents (e.g., neurotropicfactors such as NGF or BDNF).

The compositions can also be used in combination with or instead ofcompounds such as cyanoacrylates, oxidized cellulose, fibrin sealants,collagen gel, thrombin powder, microporous polysaccharide powders,clotting factors (e.g., Factor V, Factor VIII, fibrinogen, orprothrombin) and zeolite powders.

In one embodiment, vitamins may be added to the material such as vitaminK after liver surgery. In addition, other vitamins can be added tofacilitate the reconstruction of tissue or skin when applied topicallyin combination with the material. This could be after injury or in thenormal course of topical hydration.

The one or more therapeutic, diagnostic and/or prophylactic agents canbe administered simultaneously with the self-assembling materials in thesame formulation, administered simultaneously in separate formulations,or sequentially. Alternatively, the active agent(s) can be covalentlycoupled to the self-assembling material.

It will be understood that therapeutic molecules are generallyadministered in an effective amount in order to achieve a clinicallysignificant result, and effective dosages and concentrations are knownin the art. These dosages and concentrations can guide the selection ofdosages and concentrations in the present context. Bioactive moleculescan be provided at a variety of suitable concentrations and in suitableamounts (e.g., in the microgram or milligram range, or greater). Forguidance, one can consult texts such as Goodman and Gilman's ThePharmacological Basis of Therapeutics, 10th Ed., and Katzung, Basic andClinical Pharmacology.

Cells

Where cells are delivered to a patient (e.g., to promote tissuehealing), utologous cells can be used. In one embodiment, the cellscould be hematopoietic cells from the patient, dispersed in the materialand implanted. In another embodiment, the cells can be cord red bloodcells.

Molded scaffolds as described above, liquid compositions, gels, solids(e.g. powders) or other semi-solid embodiments may include one or moreadditional substances such as bioactive molecules or cells. In someinstances, the cell may secrete the bioactive molecule either naturallyor following genetic engineering (e.g., to express and/or secrete arecombinant protein). The structures described herein are able tosupport cell attachment, viability, and growth; these have been observedwhen cells are cultured on the surface of the material or when cellsgrow within the material (e.g., when encapsulated). In addition, thestructures are able to serve as substrates for neurite growth andsynapse formation when neurons are grown on or within them. Thus,bioactive molecules and cells can be encapsulated within the peptidestructures and maintain substantial function and viability when soencapsulated (see, e.g., U.S. Ser. Nos. 09/778,200 and 10/196,942).

C. Formulations

In the preferred embodiment, the formulation is a liquid orreconstitutable powder, applied topically. In one embodiment, theformulation is provided as a dry or lyophilized powder which can beadministered directly as a powder which hydrates at the site ofapplication, or suspended or dissolved in a liquid, most preferablyaqueous, and applied as a spray, paint, or injection or a hydrogel suchas chitin, collagen, alginate, or synthetic polymer. In anotherembodiment, the formulation is administered as a compressed wafer, disc,or tablet. In still another embodiment, the formulation is provided as acoating on a device, for example a stent or a catheter, which may bedissolved in an aqueous solution and dried on the device, or mixed witha polymeric carrier and applied to the device. In yet anotherembodiment, the formulation is provided in a bandage, foam or matrix, inwhich the peptides may be dispersed or absorbed. The formulation couldalso be in the form of sutures, tape, or adhesive.

Conventionally, local anesthetics are delivered by topicaladministration(e.g., formulated as an ointment, cream, or solution) orinjected into an area where the nerve fibers one wishes to block reside.The formulation may be administered to a burn or ulcer, especially whenformulated with anesthetics, antiinflammatories, growth factors, andantiinfectives, in the form of a foam, matrix or bandage, to stopbleeding or loss of interstitial fluid.

One or more of the compositions described herein can be assembled inkits, together with instructions for use. For example, the kits caninclude a biocompatible composition including self-assembling peptides(or a concentrated solution or powdered formulation thereof, togetherwith a diluent) and a vasoconstrictor, a coloring agent, or an analgesicor anesthetic agent and instructions for their combination (if notalready combined) and use (e.g., dilution and administration). The kitscan further include one or more of the additional agents describedherein. These agents can be present within a peptide-based compositionor packaged separately, and they can include one or more types ofbiological cells, an antibiotic or other therapeutic, collagen, ananti-inflammatory agent, a growth factor, or a nutrient. The kit mayalso include one or more of a syringe (e.g., a barrel syringe or a bulbsyringe), a needle, a pipette, gauze, sponges, or cotton, swabs, abandage, a nosebleed plug, a disinfectant, surgical thread, scissors, ascalpel, a sterile fluid, a spray canister, including those in which aliquid solution is sprayed through a simple hand pump, a sterilecontainer, or disposable gloves.

The formulation can be administered as appropriate for treatment of oneor more disorders. For example, the formulation may be applied to repairan injury or dealing surgery of the lung or dura, or following anepidural or spinal tap, to stop leakage of cerebrospinal fluid. Theformulation may be dispersed in a suture or adhesive for administrationat the time of or as released following suturing or gluing of a wound,thereby limiting bleeding, loss of tissue fluids, or other fluids suchas those produced by parenchymal tissues such as the liver, pancreas,and gastrointestinal tract. The formulation may be applied to any siteof bleeding, in a bandage, gauze, sponge, or other material, forimmediate control of bleeding, or released later to control bleeding ifthe initial treatment such as suturing or pressure is insufficient.Dried fabric, dehydrated foams or hydrogels, or bandages containing theformulation may be part of first aid kids for treatment of injuries, forexample, in war, at accident sites, or clinics where rapid treatment maybe required and storage space is limited.

In some embodiments, compositions including self-assembling materialscan be associated with surgical sponges. For example, liquidcompositions can be drawn into commercially available sponges prior toor during their use. Studies indicate that hemostasis can besatisfactorily achieved without traditional sponges, but there may beinstances where including compositions containing a self-assemblingmaterial may be beneficial (e.g., where a patient is experiencingprofound bleeding or where the goal of treatment is temporarystabilization). The compositions employed can include any of thenon-fibrous agents described herein. The sponges can be any known in theart, including woven and non-woven sponges and those designedspecifically for dental or ophthalmic surgeries. See, e.g., U.S. Pat.Nos. 4,098,728; 4,211,227; 4,636,208; 5,180,375; and 6,711,879.

In embodiments featuring bandages or dressings, the bandage or dressingcan include a first layer of sufficient shape and size to cover a woundor a substantial portion thereof (e.g., the most injured portion of thetissue or the area bleeding most profusely). The first layer can have atop surface, a bottom surface, and a perimeter that is, optionally,wholly or partially covered with an adhesive. A second layer of thebandage or dressing can be detachably affixed to the bottom surface ofthe first layer, optionally excluding the perimeter or any part of theperimeter bearing adhesive, and can include a liquid or non-liquidcomposition (e.g., a gel, paste, foam, cream, ointment, or powderedcomposition) including self-assembling peptides. The composition willcome in contact with the wound upon application of the bandage ordressing and is transferable from the bandage or dressing to the woundsite upon removal of the first layer or the first and second layers. Insimpler configurations, the composition comprising self-assemblingmaterials can be associated with the bottom of the first layer (e.g.,interior to the adhesive perimeter), and the second layer can beomitted. In either case, either the first and/or second layers caninclude a transparent window, through which some or all of theunderlying wound can be viewed. The composition including theself-assembling materials can be added to the bandage before it ispackaged or just before use. In another embodiment, the formulation mayinclude a further physical barrier, such as a layer of silicon film, toprevent loss of fluid by drying, after the active flow of fluids hasbeen stopped by application of the formulation.

The formulations may also be administered as immediate or controlledrelease formulations. A delayed release dosage form is one that releasesa drug (or drugs) at a time other than promptly after administration. Anextended release dosage form is one that allows at least a twofoldreduction in dosing frequency as compared to the drug presented as aconventional dosage form (e.g. as a solution or prompt drug-releasing,conventional solid dosage form). A modified release dosage form is onefor which the drug release characteristics of time, course and/orlocation are chosen to accomplish therapeutic or convenience objectivesnot offered by conventional dosage forms such as solutions, ointments,or promptly dissolving dosage forms. Delayed release and extendedrelease dosage forms and their combinations are types of modifiedrelease dosage forms.

Matrix forming materials are materials which form strong, viscous gelsupon hydration and provide control of drug diffusion and release. Inhydrophilic matrix systems, matrix forming materials are uniformlyincorporated throughout the tablet. Upon contact with water, the outertablet layer is partially hydrated, forming a gel layer. The rate ofdiffusion of the drug(s) out of the gel layer and the rate of erosion ofthe gel layer determine overall tablet dissolution and drug deliveryrates. Examples of matrix forming materials include cellulose ethersthat are water-soluble such as methylcellulose, ethyl cellulose andhydroxypropyl methylcellulose.

Formulations are prepared using a pharmaceutically acceptable “barrier”composed of materials that are considered safe and effective and may beadministered to an individual without causing undesirable biologicalside effects or unwanted interactions. The “carrier” is all componentspresent in the pharmaceutical formulation other than the activeingredient or ingredients. The term “carrier” includes but is notlimited to diluents, binders, lubricants, disintegrators, fillers,matrix-forming compositions and coating compositions.

“Carrier” also includes all components of the coating composition whichmay include plasticizers, pigments, colorants, stabilizing agents, andglidants. The delayed release dosage formulations may be prepared asdescribed in references such as “Pharmaceutical dosage form tablets”,eds. Liberman et al. (New York, Marcel Dekker, Inc., 1989),“Remington—The science and practice of pharmacy”, 20th ed., LippincottWilliams & Wilkins, Baltimore, Md., 2000, and “Pharmaceutical dosageforms and drug delivery systems”, 6th Edition, Ansel et al., (Media,Pa.: Williams and Wilkins, 1995) which provides information on carriers,materials, equipment and processes for preparing tablets and capsulesand delayed release dosage forms of tablets, capsules, and granules.

Examples of suitable coating materials include, but are not limited to,cellulose polymers such as cellulose acetate phthalate, hydroxypropylcellulose, hydroxypropyl methylcellulose, hydroxypropyl methylcellulosephthalate and hydroxypropyl methylcellulose acetate succinate; polyvinylacetate phthalate, acrylic acid polymers and copolymers, and methacrylicresins that are commercially available under the trade name Eudragit™(Roth Pharma, Westerstadt, Germany), Zein, shellac, and polysaccharides.Additionally, the coating material may contain conventional carrierssuch as plasticizers, pigments, colorants, glidants, stabilizationagents, pore formers and surfactants. Optional pharmaceuticallyacceptable excipients present in the drug-containing tablets, beads,granules or particles include, but are not limited to, diluents,binders, lubricants, disintegrants, colorants, stabilizers, andsurfactants.

Diluents, also termed “fillers,” are typically necessary to increase thebulk of a solid dosage form so that a practical size is provided forcompression of tablets or formation of beads and granules. Suitablediluents include, but are not limited to, dicalcium phosphate dihydrate,calcium sulfate, lactose, sucrose, mannitol, sorbitol, cellulose,microcrystalline cellulose, kaolin, sodium chloride, dry starch,hydrolyzed starches, pre-gelatinized starch, silicone dioxide, titaniumoxide, magnesium aluminum silicate and powder sugar.

Binders are used to impart cohesive qualities to a solid dosageformulation, and thus ensure that a tablet or bead or granule remainsintact after the formation of the dosage forms. Suitable bindermaterials include, but are not limited to, starch, pre-gelatinizedstarch, gelatin, sugars (including sucrose, glucose, dextrose, lactoseand sorbitol), polyethylene glycol, waxes, natural and synthetic gumssuch as acacia, tragacanth, sodium alginate, cellulose, includinghydroxypropylmethylcellulose, hydroxypropylcellulose, ethylcellulose,and veegum, and synthetic polymers such as acrylic acid and methacrylicacid copolymers, methacrylic acid copolymers, methyl methacrylatecopolymers, aminoalkyl methacrylate copolymers, polyacrylicacid/polymethacrylic acid and polyvinylpyrrolidone. Some of thematerials which are suitable as binders can also be used asmatrix-forming materials such as hydroxypropyl methyl

cellulose, ethyl cellulose, and microcrystalline cellulose.

Lubricants are used to facilitate tablet manufacture. Examples ofsuitable lubricants include, but are not limited to, magnesium stearate,calcium stearate, stearic acid, glycerol behenate, polyethylene glycol,talc, and mineral oil.

Disintegrants are used to facilitate dosage form disintegration or“breakup” after administration, and generally include, but are notlimited to, starch, sodium starch glycolate, sodium carboxymethylstarch, sodium carboxymethylcellulose, hydroxypropyl cellulose,pre-gelatinized starch, clays, cellulose, alginine, gums or cross linkedpolymers, such as cross-linked PVP (Polyplasdone™ XL from GAF ChemicalCorp).

Stabilizers are used to inhibit or retard drug decomposition reactionswhich include, by way of example, oxidative reactions.

Surfactants may be anionic, cationic, amphoteric or nonionic surfaceactive agents. Suitable anionic surfactants include, but are not limitedto, those containing carboxylate, sulfonate and sulfate ions. Examplesof anionic surfactants include sodium, potassium, ammonium salts of longchain alkyl sulfonates and alkyl aryl sulfonates such as sodiumdodecylbenzene sulfonate; dialkyl sodium sulfosuccinates, such as sodiumdodecylbenzene sulfonate; dialkyl sodium sulfosuccinates, such as sodiumbis-(2-ethylthioxyl)-sulfosuccinate; and alkyl sulfates such as sodiumlauryl sulfate. Cationic surfactants include, but are not limited to,quaternary ammonium compounds such as benzalkonium chloride,benzethonium chloride, cetrimonium bromide, stearyl dimethylbenzylammonium chloride, polyoxyethylene and coconut amine. Examples ofnonionic surfactants include ethylene glycol monostearate, propyleneglycol myristate, glyceryl monostearate, glyceryl stearate,polyglyceryl-4-oleate, sorbitan acylate, sucrose acylate, PEG-150laurate, PEG-400 monolaurate, polyoxyethylene monolaurate, polysorbates,polyoxyethylene octylphenylether, PEG-1000 cetyl ether, polyoxyethylenetridecyl ether, polypropylene glycol butyl ether, Poloxamer™ 401,stearoyl monoisopropanolamide, and polyoxyethylene hydrogenated tallowamide. Examples of amphoteric surfactants include sodiumN-dodecyl-β-alanine, sodium N-lauryl-β-iminodipropionate,myristoamphoacetate, lauryl betaine and lauryl sulfobetaine.

If desired, the tablets, beads, granules or particles may also containminor amount of nontoxic auxiliary substances such as wetting oremulsifying agents, dyes, pH buffering agents, and preservatives.

In one type of formulation, the material can be utilized as a shavingcream or hand lotion additive, to form a barrier for loss of fluids andas a barrier to adhesions and contamination.

Extended release formulations are generally prepared as diffusion orosmotic systems, for example, as described in “Remington—The science andpractice of pharmacy” (20th ed., Lippincott Williams & Wilkins,Baltimore, Md., 2000). A diffusion system typically consists of twotypes of devices, a reservoir and a matrix, and is well known anddescribed in the art. The matrix devices are generally prepared bycompressing the drug with a slowly dissolving polymer carrier into atablet form. The three major types of materials used in the preparationof matrix devices are insoluble plastics, hydrophilic polymers, andfatty compounds. Plastic matrices include methyl acrylate-methylmethacrylate, polyvinyl chloride, and polyethylene. Hydrophilic polymersinclude cellulosic polymers such as methyl and ethyl cellulose,hydroxyalkylcelluloses such as hydroxypropyl-cellulose,hydroxypropylmethylcellulose, sodium carboxymethylcellulose, andCarbopol™ 934, polyethylene oxides and mixtures thereof. Fatty compoundsinclude, but are not limited to, various waxes such as carnauba wax andglyceryl tristearate and wax-type substances including hydrogenatedcastor oil or hydrogenated vegetable oil, or mixtures thereof. Incertain embodiments, the plastic material is a pharmaceuticallyacceptable acrylic polymer, including but not limited to, acrylic acidand methacrylic acid copolymers, methyl methacrylate, methylmethacrylate copolymers, ethoxyethyl methacrylates, cyanoethylmethacrylate, aminoalkyl methacrylate copolymer, poly(acrylic acid),poly(methacrylic acid), methacrylic acid alkylamine copolymerpoly(methyl methacrylate), poly(methacrylic acid)(anhydride),polymethacrylate, polyacrylamide, poly(methacrylic acid anhydride), andglycidyl methacrylate copolymers. In certain embodiments, the acrylicpolymer is comprised of one or more ammonio methacrylate copolymers.Ammonio methacrylate copolymers are well known in the art, and aredescribed in NF XVII as fully polymerized copolymers of acrylic andmethacrylic acid esters with a low content of quaternary ammoniumgroups.

Alternatively, extended release formulations can be prepared usingosmotic systems or by applying a semi-permeable coating to the dosageform. In the latter case, the desired drug release profile can beachieved by combining low permeable and high permeable coating materialsin suitable proportion.

An immediate release portion can be added to the extended release systemby means of either applying an immediate release layer on top of theextended release core using a coating or compression process or in amultiple unit system such as a capsule containing extended and immediaterelease beads. Extended release tablets containing hydrophilic polymersare prepared by techniques commonly known in the art such as directcompression, wet granulation, or dry granulation. Their formulationsusually incorporate polymers, diluents, binders, and lubricants as wellas the active pharmaceutical ingredient. The usual diluents includeinert powdered substances such as starches, powdered cellulose,especially crystalline and microcrystalline cellulose, sugars such asfructose, mannitol and sucrose, grain flours and similar edible powders.Typical diluents include, for example, various types of starch, lactose,mannitol, kaolin, calcium phosphate or sulfate, inorganic salts such assodium chloride and powdered sugar. Powdered cellulose derivatives arealso useful. Typical tablet binders include substances such as starch,gelatin and sugars such as lactose, fructose, and glucose. Natural andsynthetic gums, including acacia, alginates, methylcellulose, andpolyvinylpyrrolidone can also be used. Polyethylene glycol, hydrophilicpolymers, ethylcellulose and waxes can also serve as binders. Alubricant is necessary in a tablet formulation to prevent the tablet andpunches from sticking in the die. The lubricant is chosen from suchslippery solids as talc, magnesium and calcium stearate, stearic acidand hydrogenated vegetable oils. Extended release tablets containing waxmaterials are generally prepared using methods known in the art such asa direct blend method, a congealing method, and an aqueous dispersionmethod. In the congealing method, the drug is mixed with a wax materialand either spray-congealed or congealed and screened and processed.

The preferred coating weights for particular coating materials may bereadily determined by those skilled in the art by evaluating individualrelease profiles for tablets, beads and granules prepared with differentquantities of various coating materials. It is the combination ofmaterials, method and form of application that produce the desiredrelease characteristics, which one can determine only from the clinicalstudies. The coating composition may include conventional additives,such as plasticizers, pigments, colorants, stabilizing agents, glidants,etc. A plasticizer is normally present to reduce the fragility of thecoating, and will generally represent about 10 wt. % to 50 wt. %relative to the dry weight of the polymer. Examples of typicalplasticizers include polyethylene glycol, propylene glycol, triacetin,dimethyl phthalate, diethyl phthalate, dibutyl phthalate, dibutylsebacate, triethyl citrate, tributyl citrate, triethyl acetyl citrate,castor oil and acetylated monoglycerides. A stabilizing agent ispreferably used to stabilize particles in the dispersion. Typicalstabilizing agents are nonionic emulsifiers such as sorbitan esters,polysorbates and polyvinylpyrrolidone. Glidants are recommended toreduce sticking effects during film formation and drying, and willgenerally represent approximately 25 wt. % to 100 wt. % of the polymerweight in the coating solution. One effective glidant is talc. Otherglidants such as magnesium stearate and glycerolmonostearates may alsobe used. Pigments such as titanium dioxide may also be used. Smallquantities of an anti-foaming agent, such as a silicone (e.g.,simethicone), may also be added to the coating composition.

Polymeric Matrices

Both non-biodegradable and biodegradable matrices can be used fordelivery of the self-assembling peptides, although biodegradablematrices are preferred. These may be natural or synthetic polymers,although synthetic polymers are preferred due to the bettercharacterization of degradation and release profiles. The polymer isselected based on the period over which release is desired. In somecases linear release may be most useful, although in others a pulserelease or “bulk release” may provided more effective results. Thepolymer may be in the form of a hydrogel (typically in absorbing up toabout 90% by weight of water), and can optionally be crosslinked withmultivalent ions or polymers.

Representative synthetic polymers that can be used for delivery includepolyamides, polycarbonates, polyalkylenes, polyalkylene glycols,polyalkylene oxides, polyalkylene terepthalates, polyvinyl alcohols,polyvinyl ethers, polyvinyl esters, polyvinyl halides,polyvinylpyrrolidone, polyglycolides, polysiloxanes, polyurethanes andco-polymers thereof, alkyl cellulose, hydroxyalkyl celluloses, celluloseethers, cellulose esters, nitro celluloses, polymers of acrylic andmethacrylic esters, methyl cellulose, ethyl cellulose, hydroxypropylcellulose, hydroxy-propyl methyl cellulose, hydroxybutyl methyl

cellulose, cellulose acetate, cellulose propionate, cellulose acetatebutyrate, cellulose acetate phthalate, carboxylethyl cellulose,cellulose triacetate, cellulose sulphate sodium salt, poly(methylmethacrylate), poly(ethyl methacrylate), poly(butylmethacrylate),poly(isobutyl methacrylate), poly(hexylmethacrylate), poly(isodecylmethacrylate), poly(lauryl methacrylate), poly(phenyl methacrylate),poly(methyl acrylate), poly(isopropyl acrylate), poly(isobutylacrylate), poly(octadecyl acrylate), polyethylene, polypropylene,poly(ethylene glycol), poly(ethylene oxide), poly(ethyleneterephthalate), poly(vinyl alcohols), poly(vinyl acetate, poly vinylchloride, polystyrene and polyvinylpyrrolidone.

Examples of non-biodegradable polymers include ethylene vinyl acetate,poly(meth)acrylic acid, polyamides, copolymers and mixtures thereof.Examples of biodegradable polymers include synthetic polymers such aspolymers of lactic acid and glycolic acid, polyanhydrides,poly(ortho)esters, polyurethanes, poly(butic acid), poly(valeric acid),and poly (lactide-co-caprolactone), and natural polymers such asalginate and other polysaccharides including dextran and cellulose,collagen, chemical derivatives thereof (substitutions, additions ofchemical groups, for example, alkyl, alkylene, hydroxylations,oxidations, and other modifications routinely made by those skilled inthe art), albumin and other hydrophilic proteins, zein and otherprolamines and hydrophobic proteins, copolymers and mixtures thereof. Ingeneral, these materials degrade either by enzymatic hydrolysis orexposure to water in vivo, by surface or bulk erosion.

Bioadhesive polymers of particular interest include bioerodiblehydrogels described by H. S. Sawhney, C. P. Pathak and J. A. Hubell inMacromolecules, 1993, 26, 581-587, polyhyaluronic acids, casein,gelatin, glutin, polyanhydrides, polyacrylic acid, alginate, chitosan,poly(methyl methacrylates), poly(ethyl methacrylates),poly(butylmethacrylate), poly (isobutyl methacrylate), poly(hexylmethacrylate), poly (isodecyl methacrylate), poly (laurylmethacrylate), poly (phenyl methacrylate), poly(methyl acrylate),poly(isopropyl acrylate), poly(isobutyl acrylate), and poly(octadecylacrylate).

The matrix can be in the form of microparticles such as microspheres,where peptides are dispersed within a solid polymeric matrix ormicrocapsules, where the core is of a different material than thepolymeric shell, and the peptide is dispersed or suspended in the core,which may be liquid or solid in nature. Unless specifically definedherein, microparticles, microspheres, and microcapsules are usedinterchangeably. Alternatively, the polymer may be cast as a thin slabor film, ranging from nanometers to four centimeters, a powder producedby grinding or other standard techniques, or even a gel such as ahydrogel. The polymer can also be in the form of a coating or part of astent or catheter, vascular graft, or other prosthetic device.

The matrices can be formed by solvent evaporation, spray drying, solventextraction and other methods known to those skilled in the art.

Bioerodible microspheres can be prepared using any of the methodsdeveloped for making microspheres for drug delivery, for example, asdescribed by Mathiowitz and Langer, J. Controlled Release 5, 13-22(1987); Mathiowitz, et al., Reactive Polymers 6, 275-283 (1987); andMathiowitz, et al., J. Appl. Polymer Sci. 35, 755-774 (1988). Theselection of the method depends on the polymer selection, the size,external morphology, and crystallinity that is desired, as described,for example, by Mathiowitz, et al., Scanning Microscopy 4, 329-340(1990); Mathiowitz, et al., J. Appl. Polymer Sci. 45, 125-134 (1992);and Benita, et al., J. Pharm. Sci. 73, 1721-1724 (1984). In solventevaporation, described for example, in Mathiowitz, et al., (1990),Benita, and U.S. Pat. No. 4,272,398 to Jaffe, the polymer is dissolvedin a volatile organic solvent. The peptide either in soluble form ordispersed as fine particles, is added to the polymer solution, and themixture is suspended in an aqueous phase that contains a surface activeagent such as poly(vinyl alcohol). The resulting emulsion is stirreduntil most of the organic solvent evaporates, leaving solidmicrospheres. In general, the polymer can be dissolved in methylenechloride. Microspheres with different sizes (1-1000 microns) andmorphologies can be obtained by this method which is useful forrelatively stable polymers such as polyesters and polystyrene. However,labile polymers such as polyanhydrides may degrade due to exposure towater. For these polymers, hot melt encapsulation and solvent removalmay be preferred.

In hot melt encapsulation, the polymer is first melted and then mixedwith the solid particles of peptides. The mixture is suspended in anon-miscible solvent such as silicon oil and, with continuous stirring,heated to 5° C. above the melting point of the polymer. Once theemulsion is stabilized, it is cooled until the polymer particlessolidify. The resulting microspheres are washed by decantation withpetroleum ether to give a free-flowing powder. Microspheres withdiameters between one and 1000 microns can be obtained with this method.The external surface of spheres prepared with this technique are usuallysmooth and dense. This procedure is useful with water labile polymers,but is limited to use with polymers with molecular weights between 1000and 50000. Solvent removal was primarily designed for use withpolyanhydrides. In this method, the drug is dispersed or dissolved in asolution of a selected polymer in a volatile organic solvent likemethylene chloride. The mixture is then suspended in oil, such assilicon oil, by stirring, to form an emulsion. Within 24 hours, thesolvent diffuses into the oil phase and the emulsion droplets hardeninto solid polymer microspheres. Unlike solvent evaporation, this methodcan be used to make microspheres from polymers with high melting pointsand a wide range of molecular weights. Microspheres having a diameterbetween one and 300 microns can be obtained with this procedure. Theexternal morphology of the spheres is highly dependent on the type ofpolymer used. In spray drying, the polymer is dissolved in methylenechloride (0.04 g/ml). A known amount of active drug is suspended (ifinsoluble) or co-dissolved (if soluble) in the polymer solution. Thesolution or the dispersion is then spray-dried. Double walledmicrospheres can be prepared according to U.S. Pat. No. 4,861,627 toMathiowitz.

Hydrogel microspheres made of gel-type polymers such as alginate orpolyphosphazines or other dicarboxylic polymers can be prepared bydissolving the polymer in an aqueous solution, suspending the materialto be incorporated into the mixture, and extruding the polymer mixturethrough a microdroplet forming device, equipped with a nitrogen gas jet.The resulting microspheres fall into a slowly stirring, ionic hardeningbath, as described, for example, by Salib, et al., PharmazeutischeIndustrie 40-11A, 1230 (1978). Chitosan microspheres can be prepared bydissolving the polymer in acidic solution and crosslinking withtripolyphosphate. For example, carboxymethylcellulose (CMC) microsphereare prepared by dissolving the polymer in an acid solution andprecipitating the microspheres with lead ions. Alginate/polyethyleneimide (PEI) can be prepared to reduce the amount of carboxyl groups onthe alginate microcapsules.

Other delivery systems including films, coatings, pellets, slabs, anddevices can be fabricated using solvent or melt casting, and extrusion,as well as standard methods for making composites. The polymer can beproduced by first mixing monomers and peptides as described by Sawhney,et al., and polymerizing the monomers with UV light. The polymerizationcan be carried out in vitro as well as in vivo.

D. Devices for Administration

The liquid formulations may be provided in a syringe or pipette having abarrel containing a composition including self-assembling peptides and ameans for expelling the composition from an open tip of the syringe orpipette (e.g., a plunger or bulb). The syringe may consist of one ormore compartments, so that mixing of the self-assembling materials withone or more other agents occurs at the time of application. Thecompartments may also contain an excipient such as a material forming ahydrogel or adhesive in one compartment and the self-assemblingmaterials in the other compartment. In another embodiment, onecompartment may contain lyophilized powder or particles ofself-assembling peptides, and another compartment may contain solutionto dissolve or hydrate the peptides, or other powders to mix with theself assembling materials for dry application. The composition withinthe barrel can further include any of the non-fibrous agents describedherein (e.g., one or more of a vasoconstrictor, a coloring agent, ananesthetic or analgesic agent, an antibiotic or other therapeutic,collagen, an anti-inflammatory agent, a growth factor, or a nutrient).

The self-assembling material can be applied as a coating by spraying ordipping the device into the material, the material can be impregnatedinto a bandage, gauze or other absorbent material, the material can bemixed with a polymeric material. The material can also be formulated asa pharmaceutical foam. Pharmaceutical foams are pressurized dosage formsthat, upon valve actuation, emit a fine dispersion of liquid and/orsolid materials in a gaseous medium. In one embodiment, the foamcontains the self-assembling material, in liquid or solid form,optionally in combination with one or more active agents. Suitablepropellants include, but are not limited to, hydrofluoroalkanes (HFAs),such as 1,1,1,2-tetrafluoroethane (HFA 134a) and1,1,1,2,3,3,3-heptafluoropropane (HFA 227), hydrocarbons, and carbondioxide.

II. Methods of Administration

A. Sites of Administration

The material can be applied to a variety of different surfaces toprevent or control fluid passage or to function as a barrier. The amountof self-assembling agent is determined in part by the function of thematerial in controlling fluid flow, as well as the properties of anyother materials or structures associated with the self-assemblingmaterial, alone or in combination with other bioactive materials. Theself-assembling materials can be used to stop the movement of fluids inor out of tissues/organs.

In a first embodiment, the material is used to prevent or controlbleeding. The material may be applied as a powder, liquid, a gel, or aspart of a substrate such as a bandage or membrane. This may be appliedto a blood vessel, either within the lumen, for example at the time ofangioplasty, administered by or as a coating on a stent or catheter, orexterior to the vessel, typically at the site of anastomosis. Thematerial may be applied to tissues before, during or after surgery, toprevent bleeding, which is especially problematic with tissue such asliver, kidney or spleen, or other surgeries where there is a high riskof transfusion, or to seal and protect a tissue, for example, which isfor transplantation or reattachment. In another embodiment, theself-assembling materials can be used as a shaving cream additive wherethey can act as hemostatic agents to stop bleeding due to razor cuts, abarrier to prevent contamination of razor cuts and/or a lubricant.

The material can be used to stop the flow of fluids other than blood.The material can be applied to burns to stop leakage of interstitialfluid. The material can be applied to the dura or lung as a dural orlung sealant. In one embodiment the material can be used to repair alung after a puncture wound, thereby restoring its ability to function.

The material can also be utilized in general oral surgery,periodontistry, and general dentistry, as a barrier.

The use of the material in individuals with impaired coagulation(hemophilia, von Willebrands, vitamin K, protein S or protein Cdeficiency, 10 fulminant hepatitis, disseminated intravascularcoagulation (“DIC”), hemolytic-uremic syndrome (“HUS”)) is also animportant utility since the mechanism of action is independent of thenormal coagulation pathway. In another embodiment, the material isapplied, typically by spraying or injection, to the exterior of a tissuesuch as a tumor, to prevent breakage or metastasis at the time ofsurgery. The material controls bleeding during tumor resection, as wellas limits metastatis. This also minimizes the immune response that canbe caused by a laser during tumor resection. The material is also usefulin holding loose tumors together so that nothing is left behind whenthey are resected. There are several types of tumors that arenotoriously hard to resect because they are not held together tightly(i.e. they are not solid masses). The material is expected to beparticularly useful in tumor resection in the brain, and may be usefulin a dose-response manner for subcutaneous tumor resection. This maymake it easier to resect melanomas in the skin because it appears thatthe material also facilitates skin healing. The material can alsoinclude a marker reactive with certain types of antigens on the tumorcell surface, producing a colorimetric change to show that all of thecells have been removed or there are more that need to be resected. Theaddition of an indicator to the material as well as the ability of thematerial to act like a bio-barrier could reduce the need for second andthird operations as well as complications due to outside contaminationinto the surgical field. The material can also be used to delivermaterials such as DNA to the site of injury for an extended period oftime in vitro and for multiple treatments in vivo. Another advantage ofthe material is that it can be injected and gel in place, so that thematerial can be applied and reapplied during surgery, as necessary.

In still another embodiment, the material is particularly well suited tofunctioning as a barrier to prevent contamination, either to the tissueor from the tissue, for example, during intestinal surgery. The materialmay be applied to prepare an internal site prior to surgery, especiallysites such as the sinus cavities, and for surgeries such astransurethral and transvaginal surgery. The material should also beparticularly useful in cardiovascular surgery, where both barrier andhemostasis properties can be of value, for example, for heart valvepatients who are prone to adverse consequences such as valve ringabscesses (coat valve, add antibiotic), endocarditis (coat valve),aortic root dissection (provide immediate hemostasis).

The material in combination with a metal such as silver hasanti-adhesive properties and can inhibit angiogenesis. Accordingly, itmay be useful in decreasing scarring and adhesions. The material isapplied after surgery, or to an injury such as a burn, to decreasescarring, fluid loss, and limit infection. This has further applicationin plastic surgery, especially for protection of areas cleaned anddebrided prior to closure or skin transplant, for example, inabdominoplasty, face lifts, flap donor sites, latissimus dorsi forbreast reconstruction.

In still another embodiment, the material is administered as a slurrythat can be drunk by a patient to reduce stomach bleeding, for example,from an ulcer, or decrease acidity. Alternatively, the material could beprovided as an enema or suppository to treat hemorrhoids or to fill indiverticula. In yet another embodiment, the material can be used toprevent infertility due to adhesions in the fallopian tubes or vasdeferens.

Ans assembly is not irreversible, contained substances can be released.For example, the molecules or cells can be released from the structuresin vivo (e.g., small molecules can diffuse away and larger molecules andcells can be released as the structures degrade).

In still another embodiment, the material is used as a neuroprotectiveto minimize damage and scarring following neural injury. Peptide-basedstructures promote repair and regeneration of neural tissue (e.g., whenself-assembling peptides are applied to a lesion in the brain asdescribed in U.S. Ser. No. 10/968,790). The small size of the fiberswithin the scaffolds and/or the open “weave” structure of the materialspermits extension of cell processes and allows adequate diffusion ofnutrients and waste products in a manner that provides unique advantagesfor neural tissue regeneration.

In the course of promoting wound repair, the compositions may not onlyimprove the final outcome (e.g., reduced scar formation resulting in anoutcome that more closely resembles the original tissue), but alsoreduce the time required for healing. These results could not have beenpredicted on the basis of the results achieved following application tothe injured central nervous system, given the substantial differencesbetween neural and non-neural tissues.

Finally, the materials could be used as “nanodrapes” to prevent crosscontamination. For example, the materials could be applied as a coatingto the outside of the body and then induced to self assemble. Theself-assembled material may stop the movement of liquids into the body,the reducing the possibility of cross contamination.

B. Effective Dosages

In general, the amount of material required will vary depending onvarious factors such as the size or extent of an injury (which can, inturn, be expressed in terms of the length of an incision, the caliber ornumber of damaged blood vessels, the degree of a burn, the size anddepth of an ulcer, abrasion, or other injury). The amount may vary, forexample, from a few microliters to several milliliters or more, e.g.,tens or hundreds of milliliters. The device used to deliver the materialwill vary in accordance with the amount. For example, a syringe can beconveniently used to deliver smaller amounts, whereas a tube orsqueezable bottle would be more suitable for larger amounts. Aneffective amount (whether in reference to a scaffold, precursorsthereof, or another bioactive molecule present in the formulation),means the amount necessary to elicit an improved or desired biologicalresponse.

As will be appreciated by those of ordinary skill in this art, theeffective amount of an agent may vary depending on such factors as thedesired biological endpoint, the agent to be delivered, the nature ofthe site to which the agent is delivered, and the nature of thecondition for which the agent is administered. For example, an effectiveamount of a composition for accelerating hemostasis may be an amountsufficient to decrease the amount of blood lost between the time thatbleeding begins and the time when bleeding ends by at least 25% relativeto the amount of blood lost following treatment with cold saline or notreatment. An effective amount of a composition for acceleratinghemostasis may also be an amount sufficient to decrease the timerequired to achieve cessation of visible bleeding by at least 25%relative to the time required following treatment with cold saline or notreatment. An effective amount of a composition for promoting woundhealing may be an amount sufficient to decrease the time required toachieve a predetermined percent reduction in the size of a lesion by atleast 25% relative to the time required in the absence of suchtreatment.

The amount of the composition provided can vary depending on theseverity of the subject's condition and should be sufficient to inhibitthe unwanted movement to an extent that benefits the subject. The bodilysubstance can be blood, cerebrospinal fluid, pus, serous exudate, bile,pancreatic juice, or a substance normally contained within thegastrointestinal tract (e.g., the stomach or intestine), or urinarytract.

C. How Administered

The composition can be provided on the surface of the subject's bodyand/or provided within a cavity generated by force (e.g., by unexpectedtrauma or a surgical procedure). In this way the unwanted movement ofbodily substances can be inhibited in the context of a wide range ofsituations, including traumatic injury, a medical condition (e.g., achronic or prolonged medical condition associated with bleeding), orsurgical procedures (e.g., orthopedic surgery, dental surgery, cardiacsurgery, ophthalmic surgery, or plastic or reconstructive surgery). Forexample, where the unwanted movement of the bodily substance is theresult of trauma, the subject may have a partly or completely severedbody part, a laceration, abrasion, puncture wound, or a burn. Where thecompositions are applied to a surface of the body, they may not onlyinhibit the unwanted movement of a bodily substance, but also helpprotect the subject from contamination. For example, applying aself-assembling agent to the skin will impede the movement of anunwanted foreign substance on the skin or hair into a wound. When theunwanted movement of the bodily substance results from a chronic medicalcondition, the subject may experience recurrent bleeding. For example,the subject may be experiencing bleeding in connection with varicoseveins, including telangiectases, hemorrhoids, bleeding in the lungs(due, for example, to lung cancer, bronchitis, or a bacterial or viraldisease, including pneumonia or influenza), or esophageal varices.Medical conditions associated with recurrent bleeding can be treatedwith the compositions described herein, including those that containself-assembling peptides and a vasoconstrictor (e.g., phenylephrine,which can constitute about 0.25-0.5% of the composition). Where bleedingoccurs in the orophamyx or lungs, the compositions can be administeredthrough a metered dose inhaler. If the patient's condition hasdeteriorated to the point where artificial ventilation is required, thecompositions may be administered through a respirator or by lavage.

The unwanted movement of the bodily substance can also take place duringa surgical procedure, and that procedure can involve an incision withinthe subject's nervous system, eye, ear, nose, mouth, pharynx,respiratory system, cardiovascular system, digestive system, urinarysystem, reproductive system, musculoskeletal system, liver, orintegument. The methods can be carried out regardless of whether or notthe movement of the bodily substance was intentional. The compositionsdescribed herein can be applied before or after the unwanted movementoccurs (e.g., during a surgical procedure before the intentionaltransection of a blood vessel or after an unintentional transection of ablood vessel). For example, the surgical procedure can be carried outwith the intent to repair an aneurysm, impede bleeding within the brain,to treat esophageal varices, to treat an ulcer or to inhibit the loss ofgastric contents or intestinal contents (e.g., from a swollen orruptured appendix). The surgical procedure can involve resecting aportion of the subject's intestine. Other procedures that can be carriedout with the assistance of compositions including self-assembling agentsinclude arteriography, cardiac catheterization, insertion of a stent,assistance with a natural birth or birth by Caesarean section,hysterectomy, organ transplant, joint replacement, or excision of anintervertebral disk. These procedures are representative. The surgicalprocedure can be performed with the assistance of an endoscope orlaparoscope, and the compositions can be delivered independently or froma chamber situated within these devices and connected to a distal end bya passage for release onto the subject's tissues. Where the patient hasan ulcer, that ulcer can be an esophageal, gastric, duodenal, diabetic,or decubitus ulcer. More generally, the compositions can be applied toany disrupted area of the skin, and any of the methods described hereincan include a step of identifying a patient in need of treatment.

A self-assembling peptide nanofiber scaffold (SAPNS) can provide atransparent environment for the surgical field, while also creating anoptically clear liquid that allows operation through the resultantliquid and gel mix. The surgical field is often obscured with blood anddebris during an operation. In addition, clearing debris from thesurgical field usually requires irrigating the site with saline. Salineis only a temporary solution and needs to be continuously applied tomaintain a clear surgical field. This poses several issues: anycontamination in existence will easily spread; a small opening willrequire alternating between irrigation and operating; and duringintestinal operations use of saline can result in a massive infectionleading to post-operative complications. Using the SAPNS for biologicalconfinement will reduce post operative complications in endoscopic andopen surgical procedures. Efficacy has been demonstrated on brain,spinal cord, gastrointestinal tract, liver, muscle, arteries and veins.

For example, a partial resection is currently performed as follows. Thesurgeon performs a partial resection of the intestine to remove aprecancerous area. The incision is made and the intestines are gentlylifted out of the intraperitoneal cavity and placed on the table next tothe patient. The offending area is resected and the two ends of theintestine are then ligated together. Before the intestines are put backin the body there is a colostomy bag connected to the upper end of theintestine and the area of the operation is disinfected. The intestinesare replaced in the abdomen and are sewn back up. A drain is placed inthe abdomen to make sure there is no leakage or bleeding. In contrast,using the self-assembling peptide material, a partial resection isperformed as follows. The doctor opens the abdomen and finds theoffending part of the intestine. It is isolated with additional liquidthat is poured into the intraperitoneal cavity to isolate it from therest of intraperineal cavity. The surgeon reaches through the gel thatwas formed by the liquid and resects the intestine. The two ends areligated together and the area is checked for any changes in color.Because the gel also has an indicator die that changes to blue if thereis any leakage of gastric fluids or bacteria. All of the blue is removedwith suction. A little more material is sprayed around the area of therepair and the abdomen is sewn up.

Scar Treatment:

Experiments have demonstrated that application of the self-assemblingmaterial can be used to block formation of scarring in the centralnervous system (CNS). Administration of the materials at the site oflesion blocks the stable formation of a scar, which can permitregeneration through that site. Removing the scar that develops in thecentral nervous system (CNS) permits axons to grow across the injurysite.

Chelation Enhanced Wound Healing:

The material can be used for the delivery of a chelator such as iron toa site so it can be used by the body in the local environment to rebuildbasement membrane. In tissues that do not contain enough iron, thedelivery of iron in a stable form will help healing and the rebuildingof tissue. Metals with a cystine or cystine like residue can beincorporated in the nanomaterial so there is little or no sterichinderance with the assembly of the matrix in-vivo or in-vitro.

In summary, the self-assembling peptide material can be used to create aclean local environment to perform surgery; isolate structures andmigration of contaminates; inflate structures for surgical procedures,i.e. intestine; surround structures that are being removed that mayleak, i.e. appendix, patch holes in body; allow for surgery in dirtyenvironments; used with scope procedures to surround the organ beforethe operation to contain any leakage; used to create a barrier toprevent adhesions while performing abdominal surgery; and used for forma gasket between the scope and the insertion point of the scope.Benefits during surgery are that the material is optically clear, has along shelf life at room temperature, can be operated through it,shortens prep time, eliminates counting sponges, isolates each structurein the surgical field, shortens clean up time of the operating room,shortens surgical time, reduces or eliminates cross contamination causedby other irrigants, the material is biocompatible, the breakdownproducts are natural and are absorbed by the body. The material is easyto manipulate, can be injected at the location needed, should eliminateStaphylococcal infections, may be able to reduce the cost of surgicaltheater disposables paper, reduce biohazard bags since the material canbe boiled to sterilize after the procedure to yield steam. Since thematerial is clear it should enable the surgeon to operate faster becausethe operating field is clear of blood. The elimination of wound packingto control bleeding could reduce the operating time as much as 50% in acomplicated case. Post-op infection, due to secondary infection, may bereduced by the use of the material since it can coat the wound duringand after surgery, thus reducing contamination from foreign bodies.Post-op care may be able to use the material to reduce infection due todrainage by slowing the spread of particulate material in the abdomen orchest cavity.

While the compositions can be removed from a site of application (e.g.,a bleeding vessel) at any time, a physician may wish to allow them toremain in place even after the initial goal of promoting hemostasis hasbeen achieved in order to promote wound healing.

Where the compositions include self-assembling peptides, those peptidescan include amino acid residues that are naturally occurring and thatcan be absorbed by the body. The compositions are not difficult tomanipulate, and they can be easily dispensed on an as-needed basis.Their features (e.g., stiffness) can be altered readily by altering theconcentrations of components therein (e.g., by altering theconcentration of self-assembling peptides in a given composition). Asthe resulting, assembled structure does not significantly impair one'sview of an underlying tissue, and does not have to be removed before aprocedure can be carried out. For example, a physician can assess a burnor other surface trauma that has been treated in the field with acomposition described herein. In the operating room, a surgeon can makean initial incision through the material and can continue to operatewith standard equipment, such as scalpels and clamps, or more modernmeans, such as lasers, in an internal field to which the compositionsmay also have been applied. Another advantage may be realized in time,as use of the compositions can decrease the time required to prepare apatient for surgery. As the compositions can be applied around the siteof an incision and form a coating to protect against infectious agents,there is less need to shave a patient's skin, apply drapes, and applydisinfectants.

Given the structural integrity of the assembled scaffolds, they can beremoved from an area in which they have formed if desired. Thus, anassembled scaffold can be removed by, for example, suction, or bylifting it away with an instrument such as forceps, or wiping it awaywith a swab or gauze. For example, the scaffold can be removed afterhemostasis is achieved or in the course of cleaning a wound. Based onstudies to date, the scaffold or a majority thereof can be removedwithout damaging the underlying tissue. Where the assembled scaffoldsare formed ex vivo, they can be removed from a mold and usedsubsequently (e.g., implanted in a tissue or tissue void). Thecompositions should reduce the amount of material that requires disposalor cleaning afterward (e.g., surgical drapes, sponges, and otherbiohazards).

“Nanodrapes” can be used to replace traditional paper or cloth drapes,by limiting infection following application directly to the patient, forexample, by spraying or otherwise coating the patient or the area aroundthe surgical incision. Currently a patient is prepared for surgery byshaving, scrubbing, disinfecting and draping after positioning on thesurgical table. Then bactericide and tape is applied to the area wherethe surgery is to be performed. The self-assembling composition can beapplied in place of drapes by spraying the warm liquid onto the bodywhere it self-assembles into a thin second skin. This material has apore size that is smaller than any bacteria can fit through, so itprotects from any airborne contaminants, and because the one millimeterthick material can contain a mild anti-bactericide, that clings to thebody like a second skin. The material can also have a hydratingcomponent for the skin so it does not dry out. There is no worry aboutgetting the material into the wound site because it will be broken downby the body. Color can be added to it so it is easier to determine if itis all washed off after the operation.

A scaffold (e.g., a nanoscale structured material) can be provided byintroducing, to a subject, a precursor of the scaffold at a location, orin the vicinity of a location, where the scaffold is desired (e.g., tocontrol movement or leakage of a bodily substance, to protect a wound,or to promote tissue repair). Precursors (e.g., self-assemblingpeptides) are provided in the vicinity of a location when they areprovided at a position that is close enough to the targeted area (e.g.,a bleeding vessel, a diseased section of the digestive tract, or an areaof burned skin) that they reach the targeted area in an effectiveamount. The precursors, which may be homogenous or heterogeneous (e.g.,one may apply a single type of self-assembling peptide or a mixture oftwo or more different such peptides), can be contained within acomposition and, upon contact with physiological conditions, assemble toform the scaffold (e.g., a nanoscale structured material). Thus, theprecursors can assemble in situ (i.e., within the body of a subject ator in the vicinity of administration).

The nanoscale structured material may include, or its assembly mayinvolve, additional components present in situ, (e.g., ions). Thus,precursors such as self-assembling peptides can be applied in a solutionthat is substantially free of ions (e.g., substantially free ofmonovalent cations) and self-assemble to form a macroscopic structurewhen they come in contact with such ions in the body (e.g., in a bodilysubstance such as blood, gastrointestinal contents, and the like). Forexample, a solution containing precursors can be applied at, or in thevicinity of, a site of gastric or intestinal perforation or a site wherea surgical incision has been or will be made.

The scaffold can also be provided in the form of a gel, as theprecursors (e.g., self-assembling peptides) can be assembled prior tointroducing a composition to a targeted area (e.g., the site at which anincision will be made for a surgical procedure). The assembled structuremay assume any convenient shape.

The scaffold can also be provided by providing precursors in the form ofa dry powder. A “dry” powder will have a relatively low liquid content(e.g., sufficiently low that the particles therein are readilydispersible). Self-assembling peptides provided in the form of a drypowder will assemble when they come into contact with a bodily fluidcontaining monovalent cations, and a solution containing such ions maybe added if desired to alter the rate at which the scaffold forms or itsstiffness. Self-assembling peptides may be provided as emulsions or, asdescribed above, molded into preformed shapes that can be inserted intoa body cavity or wound site in a manner similar to the manner thatsurgical sponges are currently used. If desired, a binder can be addedto a dry powder which is then formed into a desired shape. Regardless ofthe precise manner in which the scaffold is assembled (e.g., whether bybringing a liquid formulation containing precursors into contact withthe body or a dry powder into contact with an ion-containing solution exvivo), the formed scaffolds can assume a desired shape. Where the sizeand shape is such that the scaffold fills the lumen of a blood vessel,the scaffold can be used a vascular plug.

A preventative measure can be carried out before a subject experiencesan unwanted event (e.g., before an injury occurs or before bleedingbegins). Thus, the site of administration can be a site of potentialmovement or potential leakage, and the application can be made toprevent or minimize such movement or leakage should it occur. When usedin the context of a therapeutic procedure or treatment, the compositionscan reverse, alleviate, or inhibit the progress of a condition (e.g., astate, syndrome, disease, or a sign, symptom, or manifestation of such).Methods of treating a subject are generally carried out once the subjectis recognized as having a condition amenable to treatment, and any ofthe methods described herein, whether best described as prophylactic ortherapeutic, can include a step of identifying an amenable subject.

As the compositions described here can be used to inhibit movement of abodily substance in a subject, including movement within or from theepidermis, the compositions can be employed in the context of performingsurgery and may be described as new methods for performing surgery orgenerating a surgical field. The methods, whether performed in thecontext of surgery or not, can include a step of identifying a subjectin need of treatment and a step of providing a nanoscale structuredmaterial, or a precursor thereof, at or in the vicinity of a site whereunwanted movement has occurred or is expected to occur. For example, onecan identify a patient who is about to undergo a surgical procedure andprovide a biocompatible composition comprising self-assembling peptidesand a vasoconstrictor, a coloring agent, or a local anesthetic agent toa site at which an incision or other invasive maneuver will be made orhas been made. The bodily substance that is affected may be a fluid suchas blood or a blood product, serous exudate (an inflammation-associatedexudate composed largely of plasma, which typically appears as a clearor amber-colored fluid), pus, gastric juice, urine, bile, cerebrospinalfluid (CSF), pancreatic juice, and the like. The bodily substance may beviscous, sludge-like or semi-solid but will generally exhibit an abilityto flow or move. Substances of this nature include the contents of thegastrointestinal tract. The composition may be removed after application(e.g., after hemostasis is achieved or an operation on the bowel iscomplete) or may be left in place. For example, the compositions can beapplied to accelerate hemostasis or inhibit movement of intestinalcontents during surgery and some or all of the scaffold may be left inplace when the operation is complete. This provides a substantialadvantage relative to the use of sponges and other materials that mustbe removed prior to closure. The compositions can be removed in avariety of ways (e.g., by wiping or by suction).

The compositions can also be applied to shield an underlying area (e.g.,an area of burned or otherwise injured skin or other tissue) and can,therefore, help to prevent contaminants (e.g., foreign substances) fromcoming into contact with the area (i.e., the compositions can be used asa barrier or shield). A physician or other health-care provider canexamine a wound through the material, and a surgeon can operate throughit, while it is in place. Contaminating substances that have landed onthe material during the procedure could then be removed by virtue ofremoving the material.

The compositions can be administered to stabilize a wound prior todefinitive treatment (e.g., while the victim is awaiting transport to ahospital or during transit). The compositions are similarly useful whereoperations are conducted under conditions of less than optimal sterility(e.g., in field hospitals or in areas of the world where access tosterile operating rooms is limited). The compositions and methods havethe potential to significantly reduce the likelihood of contamination ininstances such as these.

The self-assembling peptide material can also be locally applied incombination with anesthetic in the local area where a procedure is totake place and can be applied at a higher concentration to reduce organmovement during surgery. This may reduce cognitive deficits to olderpatients by reducing the general anesthetic load. A thin layer can besprayed on the tissue or skin where the surgeon is operating. It can beapplied separately or together, administering specific anesthetic forspecific organs. Skin has different receptors than intestines and theneed for a specific anesthetic is needed for each of the organs.Intestines need to stop moving during surgery while the blood and bloodvessel contraction need to remain constant.

Treatment and Prevention of Bleeding:

Any individual who has an increased risk of suffering undesirablebleeding, which may or may not be excessive or immediatelylife-threatening, can be treated with the compositions described herein.These individuals include those with blood clotting disorders such ashemophilia, patients who are receiving anticoagulant therapy, patientswho suffer recurrent nosebleeds, and individuals undergoing surgery,particularly major surgery or procedures that involve accessing anartery. Without limitation, the surgery or procedure can be an operationon the nervous system, eye, ear, nose, mouth, pharynx, respiratorysystem, cardiovascular system, digestive system, urinary system,musculoskeletal system, integumentary (skin) system, or reproductivesystem. Specific examples of surgeries and procedures in which thecompositions can be used include arteriography, angiocardiography,cardiac catheterization, repair of obstetric laceration, removal ofcoronary artery obstruction, insertion of stent, Caesarean section,hysterectomy, reduction of fracture, coronary artery bypass graft,cholecystectomy, organ transplant, total joint (e.g., knee, hip, ankle,shoulder) replacement, appendectomy, excision or destruction ofintervertebral disk, partial excision of the large intestine,mastectomy, or prostatectomy.

Accident victims, individuals engaged in combat, and women giving birthare also at risk of experiencing significant blood loss. Thecompositions can be applied to a site of obstetric bleeding (e.g.,within the uterus, vagina, or neighboring tissue) in order to acceleratehemostasis. For example, the compositions can be applied to a placentaltear or used to pack the uterus to control bleeding. As with otherindications, compositions applied to the reproductive tract can beremoved or left in place. Spontaneous hemorrhage, aneurysm rupture,esophageal varices, gastric ulcers, ulcers of the upper portion of theintestine (e.g., duodenal ulcers) are also medical conditions in whichconsiderable bleeding can occur, and these individuals can also betreated as described here.

The precise source of the bleeding can vary and can be from any bloodvessel in the arterial or venous system (e.g., an artery, arteriole,capillary or capillary bed, venule, or vein). The size of the vessel mayrange from large (e.g., the compositions can inhibit bleeding from theaorta, the iliac or femoral artery, or a portal vein) to small (e.g., acapillary), and the vessel may be located anywhere in the body (e.g., ina solid organ such as liver, the stomach, intestine, skin, muscle, bone,the lungs, or the reproductive system).

The time normally required for blood clotting can be prolonged whenplasma levels of clotting factors and/or platelets are low or in casesin which an individual has received an anticoagulant (e.g., warfarin orheparin). Bleeding frequently persists for considerably longer than theaverage clotting time when there is more than minimal damage to bloodvessel integrity. Based on the studies, it is expected that thecompositions will cause hemostasis in a period of time that is lessthan, and in at least some cases much less than, the average bloodclotting time. Although the compositions are not limited to those thatachieve hemostasis in any given time (and uses such as protecting anarea from contamination or promoting tissue healing are independent ofthis function), the compositions may confer a benefit to a bleedingsubject in as little as five seconds following application. Othercompositions can exert an effect in about 10, 15, or 20 secondsfollowing application. The effective period can be characterized in amanner other than absolute time. For example, compositions may reducethe time required to achieve hemostasis by between 25% and 50%; between50% and 75%; or between 75% and 100% relative to the time required wheniced saline is applied. The time required to achieve hemostasis can bereduced by approximately 2-, 3-, 4-, or 5-fold relative to the timerequired when iced saline is applied.

The peptide concentration may be selected with reference to variablessuch as the caliber of the vessel, the extent to which it has beeninjured, and the force with which blood is exiting (or would exit uponinjury). Higher peptide concentrations will be desirable to promotehemostasis from a major vessel (e.g., the aorta, brachiocephalic,carotid, subclavian, celiac, superior mesenteric, renal, iliac, femoral,or popliteal arteries). Useful concentrations can range from betweenapproximately 0.1-10% (e.g., 1-10%; 0.5-5%; 14%; 0.1-2%; 0.1-3%; 0.1-4%;0.1-5%; and 1-8% (e.g., about 1, 1.5, 2, 2.5, 3, 4, 5, 6, or 7%). Anysubrange, or any specific value within any of the aforesaid ranges, canbe used. Any of the aforementioned concentrations may also be used forthe other indications described herein.

As noted, bleeding can be due to any of a large number of differentcauses and can be internal or external. The compositions can be appliedregardless of the cause or the nature of the cause (e.g. whether causedby a disease process or intentional or accidental trauma). Thecompositions can be used to achieve hemostasis in a confined space(e.g., inside a hollow organ) or at or near the body's surface. Forexample, the compositions can be applied to a partly or completelysevered body part such as a limb or digit. In that event, thecompositions may be serving multiple functions; they may not onlypromote hemostasis, but also protect the wounded tissue fromcontaminants and promote tissue healing. More specifically, thecompositions can be applied to a wound, left in place for a period oftime sufficient to achieve hemostasis and for blood clotting to occur,and then removed. Contaminating material such as particulates andinfectious agents adhered to the peptide gel would be removed with it. Asterile dressing may then be applied. Of course the compositions can beapplied for purposes of cleaning a wound, preventing contamination, orpromoting tissue healing even after hemostasis has been achieved or insituations in which acceleration of hemostasis is not needed.

When used to treat a nosebleed, the compositions arc inserted into theappropriate nostril and can be left in place until the bleeding hassubsided. The compositions can be easily removed by suction (e.g., usingan eyedropper or syringe) or may be removed by other physical means,including simply blowing the nose.

The compositions can also be left in place on a wound, and a dressingcan be applied over the composition. Since the composition itself iseasily removed, its presence under the dressing can help prevent thedressing from sticking to the damaged tissue. If desired, a bandagehaving a transparent portion may be used so the injured site can beviewed through the transparent portion of the bandage and the peptidestructure below. This would allow a physician to monitor the progress ofthe healing without removing the dressing. Modified bandages aredescribed further below and are within the scope of the presentinvention.

Many medical procedures involve vascular puncture, which can be followedby significant bleeding. A self-assembling peptide composition can beapplied to the wall of a punctured vessel, e.g., during withdrawal of aninstrument used to puncture the vessel. A vascular plug formed fromself-assembling peptides provides an alternative to existing vascularplugs and devices such as those described in U.S. Pat. Nos. 5,192,302;5,222,974; 5,645,565; and 6,663,655. The vascular plug can be formed insitu (e.g., at a site of vascular puncture), or can be preformed andapplied to the site.

More generally, compositions comprising nanostructured materials orprecursors thereof (e.g., self-assembling peptides) can be used forsealing any passage through tissue. The present methods thereforeinclude methods of sealing a passage through tissue by applying acomposition comprising a nanoscale structured material (e.g.,self-assembling amphiphilic peptides) to one or both ends of the passageor to its interior. The tissue can be, for example, the wall of a bloodvessel, the wall of an organ, subcutaneous tissue, or adipose tissue.Sealing the passage can result in hemostasis. The passage can also be afistula (i.e., an abnormal connection between two organs or bodystructures or between an organ or structure and the external world). Ifdesired, a surgeon can apply the compositions to the interior of atubular structure such as the intestine or a blood vessel, resect andligate the intestine or blood vessel in the gel, and evacuate the gelfrom the interior of the structure to restore continuity of thestructure and allow reperfusion of the area with blood or other bodysubstances. The materials may also be used to limit reperfusion injury.For example, the self-assembling materials can be administered postischemia, such as to patients who have been treated with a thrombolyticagent. The materials may also be used to limit reperfusion injurythrough the re-establishment of blood tissue barriers prior to, during,and/or after reperfusion. For example, the self-assembling material maybe used to re-establish blood tissue barrier through the internalcoating of portions of the circulatory system. This may be beneficial indiseases such as ischemic infarction, hemorrhagic stroke, or reperfusioninjury. Finally, the self-assembling materials may be used to limitreperfusion injury through the re-establishment of the integrity of thevascular structure prior to, during, and/or after reperfusion.

For surgical applications, the wound or any part of the surgical fieldcan be packed with a composition comprising self-assembling peptides.This approach can be used instead of wound packing as it isconventionally performed during surgery. As the compositions containbiocompatible and biodegradable material, they can be left in place,thereby avoiding the need for removal at the end of the procedure andavoiding the need for a subsequent operation for this purpose.Biodegradable materials can be broken down physically and/or chemicallywithin cells or within the body of a subject (e.g., by hydrolysis underphysiological conditions or by natural biological processes such as theaction of enzymes present within cells or within the body) to formsmaller chemical species which can be metabolized and, optionally,reused, and/or excreted or otherwise disposed of. Preferably, thebiodegradable compounds are biocompatible.

Gastrointestinal bleeding, which can occur as a consequence of ulcers orangiodysplasia, is a relatively common and serious condition that can befatal if left untreated. Bleeding esophageal varices, and bleedinggastric or duodenal ulcers can be particularly severe. A number ofendoscopic therapeutic approaches have been developed to achievehemostasis, such as the injection of sclerosing agents, the attachmentof mechanical hemostatic devices, and contact electrocautery techniques.The compositions can be administered at, or in the vicinity of, an ulceror a site of bleeding in the esophagus, stomach, small intestine, orlarge intestine. Bleeding in the distal portion of the large intestine,rectum, or anus (e.g., hemorrhoids) can also be treated in this manner.

Rupture of an aneurysm can represent a catastrophic event with rapidlyfatal consequences. Ruptured aortic aneurysms can rapidly result inexsanguination despite prompt medical attention. Ruptured intracranialaneurysms frequently have devastating consequences. The compositions andmethods of the invention can be used to treat bleeding from a rupturedaneurysm in an essentially similar manner to the way in which they areused to treat bleeding due to other causes (e.g., by application ofself-assembling precursors or a preformed structure to the site ofbleeding). Given the often severe consequences of aneurysm rupture,surgical repair is often attempted. The compositions can be applied inthe context of any attempted repair (e.g., during open surgery orendovascular repair (e.g., with placement of a graft and/or stent)).More specifically, the present methods include treating an aneurysm byintroducing a composition comprising a nanoscale structured material orprecursor thereof (e.g., a composition comprising self-assemblingpeptides) into the aneurysm (e.g., into the aneurysm sac). Once anybleeding is under better control, the aneurysm may then be repairedusing any suitable technique. Presence of the peptide structure withinthe aneurysm sac reduces the chance of leakage or rupture prior to orduring these other procedures. The scaffold can be left in place.

Inhibiting Movement or Leakage of Cerebrospinal Fluid (CSF):

The dura mater is the tough, outermost, fibrous membrane that covers thebrain and spinal cord, and lines the inner surface of the skull. Leakageof CSF is a significant complication following injury, surgery, or otherprocedures in which the dura mater is penetrated, including inadvertentpenetration in the course of administering an anesthetic to the epiduralspace. Such leakage can lead to serious sequelae, such as severeheadaches, infection, and meningitis. The composition can inhibitmovement or leakage of CSF in a subject in need thereof afterapplication at, or in the vicinity of, a site of unwanted movement orleakage of CSF. The compositions can be applied over sutures followingdura mater surgery to help prevent CSF from leaking out of the incisionsite. The compositions can also be used to inhibit movement or leakageof fluid from the ear drum.

Inhibiting Leakage of Contents of the Gastrointestinal Tract:

The compositions can inhibit the movement of gastrointestinal contents.For example, the structures can prevent leakage of gastrointestinalcontents following gastric or intestinal perforation or during surgery(see Example 4). The structures can be used to isolate such bodilysubstances and prevent their spread within the peritoneal cavity,thereby minimizing contamination and the risk of subsequent chemicalperitonitis and/or infection. Gastric contents, which contain digestivesecretions of the stomach glands consisting chiefly of hydrochloricacid, mucin, and enzymes such as pepsin and lipase, can cause injuryand/or infection if released into the peritoneal cavity. Release ofintestinal contents into the peritoneal cavity represents a frequentevent during surgery on the intestine and can also occur in cases ofintestinal perforation or a ruptured appendix. The composition can beused to inhibit leakage of gastrointestinal contents into the peritonealcavity. The site of movement can be a site of gastric or intestinaldamage caused by a disease process or a surgical incision. Thecompositions can be applied to the exterior of any organ in thedigestive system (e.g., the stomach, or small or large intestine) or canbe injected or otherwise introduced into their interior. Thecompositions can be administered in the course of resecting a segment ofthe intestine. For example, one can fill a segment of intestine thatextends from a first point to a second point with a present compositionand resect a portion of the intestine that lies between the first andsecond points. In one embodiment, the self-assembling material may beused to treat heartburn. For example, the self-assembling materials canbe formulated as a solution, suspension, or emulsion (such as a drink orshake), gel, tablet, wafer, capsule, etc. that is administered orally inorder to coat portions of the gastrointestinal tract. The formulationscan be used to: stop the movement of bodily fluids including, gastricjuices and blood; coat the GI tract, and/or stop the progression ofulcers, erosion, and inflammation. The formulations may be used toprevent damage to the esophagus from acid reflux disease. Theformulations may also used to help the repair of cells in the esophagusthat were damaged by acid reflux, other diseases or disorders, and/ortherapeutic interventions. The formulations may be used to help therepair of primary and secondary ulcers and erosions to the mucosa. Theformulations may be used to deliver therapeutic, prophylactic, and/ordiagnostic agents to portions of the GI tract as needed. For example,the self-assembling materials may be used to deliver agents tore-establish the flora and fauna of the GI tract which have been deletedto radiation treatment and/or disease or trauma.

In a related method, one can use the compositions to remove intestinalcontents that have been released into the peritoneal cavity. The methodincludes applying a liquid composition to the released intestinalcontents, allowing the liquid composition to undergo a phase transition,and then removing the gel-like or semi-solid composition. These stepscan be repeated once or more until the surgeon is satisfied with theamount of intestinal contents that have been removed from the peritonealcavity.

One can similarly inhibit movement of the contents of other internalorgans (e.g., organs in the biliary or urinary systems). For example,one can inhibit movement of bile, pancreatic juice (i.e., secretions ofthe exocrine portion of the pancreas that contain digestive enzymes), orurine and/or decontaminate or clean an area into which bile, pancreaticjuice, or urine have been released by application and subsequent removalof the compositions to the site. The methods thus have broad applicationto surgeries for repairing or otherwise treating intestinal, biliary,and/or urinary system defects.

Wound Healing:

Studies also indicate that the compositions have the ability to enhancehealing, particularly of an epithelial layer or muscle, and cantherefore be administered to treat a site of tissue damage. For example,one can apply a composition including self-assembling peptides to thesite of tissue damage. The compositions appear to both increase the rateof tissue repair and inhibit formation of scar tissue. The compositionscan be used for either acute or chronic wound care. For example, theycan be applied to skin wounded in any manner (e.g., lacerated or burned)and to lesions such as diabetic ulcers and pressure sores.

These materials can be used to maintain hydration and nutrition topatients that have had burns or in cases the outer skin has beenbreached due to abrasion or burn.

In another case the material can be used to maintain body temperaturewhen the patient is covered with the material by means of external heator cooling source.

Tissue Regeneration

Drug Delivery Vehicle to the Intrathecal Space:

The materials described herein may be used to deliver therapeutic and/orimaging agents to the intrathecal space. Examples of therapeutic agentsinclude, but are not limited to, anti-inflammatory agents and agents tostimulate nerve/spinal cord regeneration. Hydrogel material have beenused to attempt to delivery of one or more active agents to theintrathecal space. However, these materials can be limited by their slowpolymerization times, which allows for the material to diffuse awaybefore the material polymerizes to form the gel. The materials describedherein can be designed to self-assembly quickly so that the materialdoes not diffuse away.

Cartilage Repair:

The materials described herein may be used for cartilage repair. Thematerials would typically be injected into the site where cartilagerepair is needed. The material can used alone or in combination withcells and/or growth factors.

Bone Regeneration:

The materials described herein may be used to prepare compositematerials for bone regeneration. For example, the self-assemblingmaterials can act as a carrier for inorganic materials, such as calciumphosphate or hydroxyapatite, organic materials, such as growth factors,and/or bone grafts. Inorganic materials such as calcium phosphate can beremodeled by the osteoclast resorption mechanism to regenerate bone. Thematerials may also be injected under the periosteum to stimulate bonegrowth as a means for creating bone grafts in vivo. Alternatively, thematerials described may be used for guided bone regeneration therapieswhich limit fibrous in growth. For example, the materials describedherein may be used in dental procedures as molds which are placed overthe tooth socket to prevent fibrous tissues from growing into the socketspace.

Oxygen Delivery:

The self-assembling materials described herein may also be used todeliver oxygen to the lungs and/or other organs. For example, thematerials can be superoxygenated to provide oxygen support/perfusion forpatients suffering from pulmonary hemorrhage and other lung diseases.

Delivery Methods, Devices, and Kits:

A variety of devices can be used to introduce the compositions to atarget area of the body. The devices can be simple, such as a syringe,and such devices can be provided together with the compositions in kits.The composition can be locally delivered at or near a target area in thebody by injection (e.g., using a needle and syringe), or with acatheter, cannula, or by dispensing (e.g., pouring) from anysuitably-sized vessel. The compositions can be delivered with theassistance of imaging guidance (e.g., stereotactic guidance) ifnecessary. Alternately, a material can be wetted with the compositionand then used to apply a composition to an area of tissue.

For storage and shipping, self-assembling materials can be dissolved ina suitable solvent (e.g., an aqueous medium such as sterile water, andstored for long periods of time prior to use). Peptide-containingsolutions have been stored for up to two years without substantial lossof activity. If partial self-assembly occurs after a prolonged period oftime, physical agitation (e.g., sonication) can be used to restore thematerial to a more liquid state prior to administration. Alternatively,the material can be applied as a gel. If desired, a small amount of ions(e.g., monovalent cations) can be added to a solution prior toapplication. This may speed the process of gel formation. Alternately,monovalent cations can be applied after the solution has beenadministered.

Kits containing syringes of various capacities or vessels withdeformable sides (e.g., plastic vessels or plastic-sided vessels) thatcan be squeezed to force a liquid composition out of an orifice areprovided. In one embodiment, the syringe or vessel contains multiplecompartments, one containing monovalent ions, and the otherself-assembling peptides, which are mixed at the time of administration,through a common needle. An endoscope can be used to deliver thecompositions for treatment of a hollow organ (e.g., the esophagus,stomach, intestine, etc.) or body cavity (e.g., during minimallyinvasive surgery). Minimally invasive surgery refers to an approach tosurgery whereby operations are performed with specialized instrumentsdesigned to be inserted through small incisions or natural bodyopenings, often performed with endoscopic visualization. Examplesinclude laparoscopic surgery, arthroscopic surgery, and endovascularsurgery. An endoscope is typically a long, flexible tube-like device. Inaddition to allowing visualization of internal structures, manyendoscopes have additional diagnostic (e.g. biopsy) and therapeuticcapabilities (e.g. delivery of therapeutic agents) through specialchannels. Colonoscopes, sigmoidoscopes, bronchoscopes, cystoscopes, andlaparoscopes, are variants of an endoscope having features making themparticularly well suited for viewing certain organs, structures, orcavities. Any of these devices can be used to deliver the compositions.Kits may be packaged including an endoscope and a vessel containing asolution comprising self-assembling peptides. Suitable endoscopes areknown in the art and are widely available. Endoscopes are currently inuse to deliver sclerosing agents to sites of esophageal bleeding.

Kits can include self-assembling peptides and one or more of: a syringe,a needle, thread, gauze, a bandage, a disinfectant, an antibiotic, alocal anesthetic, an analgesic agent, surgical thread, scissors, ascalpel, a sterile fluid, and a sterile vessel. The peptides can be insolution or dry (e.g., as a drypowder). Components of the kit may bepackaged individually and are sterile. The kits are generally providedin a container, e.g., a plastic, cardboard, or metal container suitablefor commercial sale. The kit may be styled as a “first aid kit,” inwhich case it will typically have a symbol such as a red cross on theexterior. Any of the kits can include instructions for use.

EXAMPLES Example 1: Self-Assembling Peptide Material AcceleratesHemostasis in the Brain

Complete transection of a branch of the superior sagittal sinus in thebrains of rats and hamsters was performed after removing a portion ofthe skull overlying the transected tissue. Animals were anesthetizedwith an i.p. injection of ketamine (80 mg/kg) and xylazine (8 mg/kg).All surgical procedures were conducted under an operating microscope.Twenty-two animals, including 10 adult hamsters and 12 young adultfemale Spraque-Dawley rats (200-250 g), were treated with either icedsaline or 20 μl of a 1% peptide solution at the site of the sinus branchtransection. The material was prepared by dissolving RADA 16-1(n-RADARADARADARADA-c; SEQ ID NO: 1) peptide in sterile water, and thepeptide-containing solution was applied to the injured tissue with a 31gauge needle attached to a 2 cc syringe.

The experiment was videotaped with a time stamp and was replayed oneframe at a time to evaluate the length of time required for the peptidesolution to form a gel, which effectively impeded bleeding. Hemostasiswas assessed visually, and “complete hemostasis” was defined as thecomplete lack of movement of blood from the wound site. Completehemostasis was achieved within 10 seconds of the application of thepeptide solution in all cases.

A series of pictures was taken of an adult rat in which a portion of theoverlying skull was removed and one of the veins of the superiorsagittal sinus was transected and then treated with a peptide-containingsolution. The initial picture shows the exposed brain and veins of thesuperior sagittal sinus; the next picture shows the cutting of the vein;the next picture shows bleeding from the ruptured vein; and the finalpicture shows the same area five seconds after the peptide solution wasapplied. Complete hemostasis was achieved.

Durations were measured from the start of application of peptidesolution to the completion of hemostasis after transection of the veinsleading to the sinus in the brains of adult rats. Complete hemostasiswas achieved in an average of 8.3 seconds. In the saline controls,cessation of bleeding was never achieved. The saline control experimentwas terminated at the same time point in order to prevent the animalsfrom bleeding to death.

Similar results have been obtained following complete transection of thesuperior sagittal sinus. A higher concentration of peptide (e.g.,approximately 3%-4%) was used in the latter experiment in order toachieve hemostasis. The three saline control cases continued to bleedafter 20 seconds. In the control animals, the iced saline was removedand the peptide solution was applied, resulting in complete hemostasisalmost immediately.

A total of 22 rats and 64 hamsters have been subjected to experiments inwhich peptide-containing solutions effectively achieved hemostasiswithin 10 seconds following application to a site intracranial bleeding.

Example 2: Self-Assembling Peptide Material Accelerates HemostasisFollowing Femoral Artery Transection

The sciatic nerve and the adjacent femoral artery were exposed in adultrats, and the femoral artery was transected. Twelve rats were treated byapplication of 20 μl of a 1% solution of RADA 16-I (SEQ ID NO: 1)peptide to the site of transection using a glass pipette attached to asyringe body, while controls were treated by applying cold saline to thesite of transection. In all treated cases, hemostasis was achieved inless than 10 seconds. The saline control cases continued to bleed untilthe experiment was terminated at 110 seconds. In these control animals,subsequent replacement of the cold saline with the peptide solutionresulted in almost immediate achievement of complete hemostasis.

A series of pictures was taken in an adult rat in which the femoralartery was transected. In the picture taken first, the sciatic nerve andthe femoral artery are exposed. The next picture shows the cutting ofthe artery, and the next picture shows bleeding. After about fiveseconds, complete hemostasis was observed in the area of a clear gelformed by the assembled peptides in the presence of blood and plasma.The assembled material can be suctioned off the site easily if desired.Complete hemostasis was maintained for the duration of the test (1hour).

Complete hemostasis was achieved in less than 10 seconds. In the salinecontrols, hemostasis was never reached.

Muscle trauma experiments showed immediate hemostasis after 1-2 cmincisions were made in the muscle on the back of a rat. Thespinotrapezius muscles on the back of the rats were exposed and a deepcut was made in the muscle, after which 1% peptide solution (RADA16-I)(SEQ ID NO: 1) was applied in the cut. Within 10 seconds, all bleedinghad stopped. With the application of iced saline alone, control animalscontinued to bleed after 20 seconds.

This procedure was duplicated in the muscle of the hind limb(porteocaudalis and musculus tibialis cranialis) and similar resultswere obtained. Between 1% to 100% peptide (RADA 16-1) (SEQ ID NO: 1) wasapplied to limb wounds, and hemostasis was achieved in all cases.However when an artery or vein was transected 2% or higher material wasneeded to bring about hemostasis. With the application of iced salinealone, control animals continued to bleed after 20 seconds.

Example 3: Self-Assembling Peptide Material Accelerates Hemostasis inLiver

To further demonstrate the ability of peptide-containing structures tohalt bleeding of a vessel having relatively low pressure, theintraperitoneal cavity of an adult rat was opened, the liver wasexposed, and the lobus sinister lateralis received a rostral to caudalcut completely transecting a portion of the liver. Profuse bleedingensued. A 1% peptide solution (RADA16-1) (SEQ ID NO: 1) was applied tothe cut and in its vicinity using a 27 gauge needle and 4 cc syringe.All bleeding stopped within 10 seconds. A series of pictures wasobtained. The first shows exposure of the liver; in the second, theliver is separated, and profuse bleeding is evident; and in the third,the two portions of the liver are allowed to come back together, and thebleeding continues. After treating the site with 1% peptide solution(applied topically and in the cut), all bleeding stopped within 10seconds. A clear area was observed between the two halves of the lobussinister lateralis. This procedure was repeated several times with thesame result.

A similar experiment demonstrated the ability of the peptide structuresto halt bleeding of a vessel in the liver having a higher pressure. Aseries of pictures illustrate the experiment. The first depicts theopened intraperitoneal cavity and exposed liver; in the second, thelobus sinister lateralis received a transverse cut completelytransecting a portion of the liver and a major branch of the portalvein; and the third shows profuse bleeding from the site of injury. Thecut was treated with 4% peptide solution applied topically and in thecut. All bleeding stopped within 10 seconds. The lower part of the lobussinister lateralis was pulled downward to show that the peptidestructure is in the cut. The site did not bleed even when subjected tothis physical stress. Ten minutes later, there was still no bleeding.Thus, application of 4% peptide solution brings about completehemostasis in a high pressure bleeding environment in less than 10seconds.

Treatment with a 2% or 3% peptide solution was tested in the same typeof experiment and complete hemostasis was also achieved. Treatment witha 1% solution resulted in partial cessation of bleeding. In addition, 30seconds after treatment the excess peptide structure was wiped away fromthe injury site and hemostasis was maintained. This procedure wasrepeated several times with the same result.

In other experiments ¼ of the lobe in the lower right quadrant of thelobus sinistras laterialis was removed, and the margin was treated witha topical application of 2% peptide (RADA16-I) (SEQ ID NO: 1) to thesite of injury. Bleeding stopped in less than 10 seconds. One minutelater the peptide was removed, and complete hemostasis was achieved atthe margin of the liver.

Example 4: Self-Assembling Peptide Material to Prevent Adhesions

The liver of 18 adult rats was exposed under deep anesthesia, the upperright lobe was punched by a 4 mm punch, then the wound was treated with3% NHS-1. Animals were allowed to survive 2 d, 7 d, 14 d, 6 w, and 8 wrespectively, then the animals were anesthetized again and the punchedlobe of liver was dissected and processed for H&E staining. In additiona set of controls were treated with either saline or cautery.

4 mm liver punch biopsy experiment with control and filling of punchwith 3% RADA. All of the controls had adhesions on both surfaces whilethe treated had no adhesions. In the 2 week, 6 week and 8 week controlsadhesions were cut away on the upper and lower surfaces. On all the 3%RADA treated cases there were no adhesions on with the upper or lowersurface of the liver.

Example 5: Self-Assembling Peptide Material

The intestine of an adult rat was perforated with a small cut at thelevel of the duodenum that resulted in the leakage of gastric fluid intothe intraperitoneal cavity. When the site was treated with 2% peptide(RADA16-I) (SEQ ID NO: 1) solution all leakage of gastric fluids fromthe intestine stopped. An additional volume of 2% peptide solution wasinjected into the duodenum at the level of the injury. This preventedall leakage from the intestine for one hour, the duration of theprocedure. In the control cut at the level of the duodenum, the wall ofthe intestine inverted and gastric fluids continued to leak from thesite of injury when left untreated. When the site was treated withpeptide solution 15 minutes after the injury, the peptide treatment alsostopped all leakage from this injury site. In addition, the treatmentstopped the progression of the intestinal wall inversion.

Example 6: Self-Assembling Peptide Material Accelerates Healing of SkinWounds

To demonstrate the ability of the self-assembling peptides to enhancewound healing, animals were subjected to punch biopsies of the skin andsubcutaneous tissue. The regions from which the biopsies were taken wereeither treated by a single application of self-assembling peptide(RADA16-I) (SEQ ID NO: 1) solution or were left untreated. The woundswere left unbandaged. A series of pictures of a 4 mm punch biopsyhealing test in which injured animals were treated with the selfassembling peptide and compared to matching cases with no treatmentillustrates the results. The wounds were photographed on day 0, day 1,day 4, and day 7. The treated wounds healed much faster as evidenced bythe contraction of the wound site in all three punches as early asday 1. Treatment with the peptide appeared to speed healing by as muchas 5 days in some cases. In all cases, shrinkage of the wound sitehappened faster in the treated cases.

Example 7: Compositions Containing Lidocaine

RADA 16 (SEQ ID NO: 1) mixed with lidocaine and the mixture was appliedto the skin of adult rats before applying a pin prick. It is a 5% mix oflidocaine and RADA1-16 (SEQ ID NO: 1). Applied on the skin and left forthe duration of the testing. When mixed with a self-assembling peptide,the response to pin prick was muted four times longer than the responsewas muted using lidocaine alone. In addition, we applied solutions ofself-assembling peptides and lidocaine to the intestines of two ratswhile performing intestinal surgery. The solution reduced peristalsisfor the duration of the surgery with no apparent side effects to theanimals.

The foregoing description is to be understood as being representativeonly and is not intended to be limiting. Alternative systems andtechniques for making and using the compositions and devices of theinvention and for practicing the inventive methods will be apparent toone of skill in the art and are intended to be included within theaccompanying claims.

We claim:
 1. A method of limiting formation of adhesions or scars,comprising administering to a region of a subject's body at risk offorming adhesions, a self-assembling peptidomimetic formulation in anamount effective to form a barrier structure to limit the formation ofadhesions or scars in the region, wherein the self-assemblingformulation comprises peptidomimetics selected from the group consistingof α-peptides, β-peptides, γ-peptides, δ-peptides, and co-polymersthereof, wherein the α-peptide peptidomimetics are selected from thegroup consisting of N,N′ linked oligoureas, oligopyrrolinones,oxazolidin-2-ones, azatides and azapeptides.
 2. The method of claim 1,wherein the self-assembling peptidomimetic formulation comprises acopolymer of α-peptides and one or more of β-peptides, γ-peptides, orδ-peptides.
 3. The method of claim 1, further comprising providing oneor more therapeutic agents, prophylactic agents, diagnostic agents, orcells.
 4. The method of claim 1, wherein the self-assemblingpeptidomimetic formulation comprises a pharmaceutically acceptablecarrier or matrix or support material for administration onto or intothe body.
 5. The method of claim 4, wherein the self-assemblingpeptidomimetic formulation is administered as a dry powder, liquid, orincorporated into hydrogels, or sponges.
 6. The method of claim 1comprising applying the self-assembling peptidomimetic formulation to aburn.
 7. The method of claim 1 comprising applying the self-assemblingpeptidomimetic formulation to a wound or surgery site or a site ofinflammation.
 8. The method of claim 1, wherein the self-assemblingpeptidomimetic formulation is administered as a coating on a medicaldevice or implant.
 9. The method of claim 1, wherein the self-assemblingpeptidomimetic formulation is administered to the site of a surgicalprocedure at a time point selected from group consisting of before theinitiation of the surgical procedure, during the surgical procedure,after commencement of the surgical procedure, and combinations thereof.10. The method of claim 1, wherein the self-assembling peptidomimeticformulation has a concentration of Li+, Na+, K+, and Cs+ ions less than50 mM.
 11. The method of claim 1, wherein the self-assemblingpeptidomimetic formulation is administered in the form of a solution.12. The method of claim 11, wherein the concentration of self-assemblingpeptidomimetics in solution is between 1.0% and 10.0%, inclusive. 13.The method of claim 11, wherein the concentration of self-assemblingpeptidomimetics is between 2.0% and 3.0%, inclusive.
 14. The method ofclaim 11, further comprising the step of forming the solution bydissolving a powder of the self-assembling peptidomimetics with water oroil.
 15. The method of claim 1, wherein the self-assemblingpeptidomimetic formulation is administered to damaged skin or tissuethat is bleeding to form a self-assembled barrier structure which ismaintained in contact with the tissue following the cessation ofbleeding.